G Proteins:

Implications K. Manji,

Husseini

There

is mounting

evidence

that

a family

their

treatments.

available

form

offunctional implicated

psychotropic

drugs

modulate

neuronal

G proteins

the basis

affect

and the complexities of the nervous tion ofneuronal function, it seems various major psychiatric illnesses. G protein targets (AmJ Psychiatry

remains 1992;

The

may

be one

an exciting 149:746-760)

prospect

for

ne of the most exciting recent advances in neuroscience has been the elucidation of the molecular mechanisms underlying neuronal communication. Although rapidly increasing numbers of potential neurotransmitters and receptors continue to be identified, it has become clear that the translation of the extracellular signals (into a form that can be interpreted by the complex intracellular enzymatic machinery) is generally achieved in relatively few ways. Generally speaking, these transmembrane signaling systems, and the receptors that use them, can be divided into two major groups: 1 Those with relatively self-contained structures and messages that take the form of transmembrane ion fluxes. .

with

multiple

components

and

messages

that

generate various intracellular “second messengers.” The first class of receptors contain in their stable molecular complex an intrinsic ion channel. Receptors of this class include those for a number of amino acidsincluding glutamate, y-aminobutyric acid (GABA) (through the GABAA receptor), and glycine-the nicotinic acetylcholine receptor, and the 5-HT3 receptor (1). Receptors of this type have also been described as “ionotropic” and are generally composed of four or five Received May 22, 1991; revision received Nov. 6, 1991; accepted Nov. 21, 1991. From the Section on Clinical Pharmacology, Experimental Therapeutics Branch, NIMH. Address reprint requests to Dr. Manji, Section on Clinical Pharmacology, Experimental Therapeutics Branch, NIMH, Bldg. 10, Rm. 2D46, 9000 Rockville Pike, Bethesda,

MD 20892. The Hsiao

746

author wishes for their helpful

proteins

(G pro-

in the CNS,

the involvement conditions and

endowing

the neuron

in the function and/or expression of G states, and a number of currently

understanding ofthe

keys

ofthe

mechanisms

to understanding

by which

the functioning

system. Given their widespread, critical roles in the regulalikely that G proteins are involved in the pathophysiology of The development of novel, site-specific drugs with primary

O

2. Those

integration

Abnormalities ofpathophysiologic

G proteins.

activity

binding

action in the CNS, and an emphasis on psychiatric

with

ofsignal

diversity. in a variety

triphosphate

ofa vast array ofextracellular, receptor-detected effectors. The author reviews the literature dealing generation, the role of G proteins in regulating

divergence,[neurotransmiuer ofclinical ctnditions,

G proteins

with a large degree proteins have been

M.D.

ofguanosine

teins) play an obligatory role in the transduction signals across cell membranes to intracellular with G protein coupling to second messenger both the convergence and of G proteins in a variety

for Psychiatry

to thank Drs. William Z. Potter and suggestions, input, and encouragement.

John

K.

the future.

subunits surrounding a central ionic pore, which opens transiently when the transmitter binds, allowing ions to flow either into the neuron (e.g., Na, Ca2, CF) or out of it (e.g., K), thereby generating synaptic potentials. Neurotransmission of this type is very fast: ion channels open and close within milliseconds (2). Most receptors, however, do not have ionic conductance channels within their structure; rather, they regulate cellular activity by generating various second messengers. In genera!, receptors of this class do not directly interact with the various second-messengergenerating enzymes but transmit information to the appropriate “effector” by the activation of interposed “coupling proteins.” It is now clear that a family of structurally related, guanosine-triphosphate-binding proteins (or G proteins) play an obligatory role in the transduction of a vast array of extracellular, receptordetected signals across cell membranes to intracellular effectors. Indeed, Birnbaumer (3) has suggested that about 80% of all known hormones, neurotransmitters, and neuromodulators (numbering around 100) elicit cellular responses through G proteins coupled to a Variety

of

cellular

effectors,

including

adenyly!

cyclase

(frequently referred to as adenylate cyclase), phospholipase C, phospholipase A2, and several ion channels (4). Receptors coupled to G proteins include those for catecholamines, serotonin, acetylcholine, various peptides (e.g., vasopressin, substance P), and even sensory signals, such as light and various odorants (3). Receptors coupled to G proteins are prototypic “metabotropic” receptors. Activation of these receptors initiates a cas-

Am

J

Psychiatry

1 49:6,

June

1992

HUSSEINI

cade of events, usually through regulation of intracellu!ar second messengers, that lead to modification of enzymatic phosphorylation of various membrane and cytosolic proteins (enzymatic activation or inhibition), including receptors, ion channels, and possibly G proteins themselves (1 ). As might be predicted, receptors coupled to G proteins generate electrical signals much more slowly, often with a latency of signal onset of at least 30 msec (2). The ability of many G proteins to interact with multiple receptors also provides an elegant mechanism for the neuron to respond to a large number of stimuli and to regulate both the convergence and divergence of neurotransmitter action (5, 6). The high degree of complexity generated by the interactions of G-protein-coupled receptors may be one mechanism by

which

neurons

acquire

the

flexibility

necessary

for

generating the wide range of responses observed in the nervous system; this has led to the suggestion that Gprotein-coupled receptors may be involved in pathways regulating such diverse vegetative functions as mood, appetite, and wakefulness (2). Thus, understanding the mechanisms by which G proteins

modulate

to understanding of the nervous

neuronal

of second

functioning Additionally,

is one

of

the

keys

the complexities there is a growing body of evidence implicating both quantitative and qualitative G protein abnormalities in a variety of clinical disorders. Similarly, there is increasing evidence that a number of psychotropic agents exert their clinically relevant effects at postreceptor sites, including G proteins. The identification of these putative G protein targets may thus lead not only to the development of more potent, site-specific treatments but also to the localization of molecular and biochemical factors predisposing individuals to various psychiatric illnesses. This overview will explore the growing knowledge of G protein coupling to chemical and ionic second messenger generation. Specifically, I will discuss the biochemistry

the system.

activity

messenger

and

synthesis,

the

role

of

ARE

G PROTEINS?

The G proteins are a family of GTP-binding that play an obligatory role in the transduction tracellular, receptor-detected signals across membrane to various intracellular effectors These proteins have been the focus of extensive since

the

cyclase

(the

demonstration enzyme

that which

stimulation

generates

both hormones and GTP. demonstrated that -adrenergic GTPase activity (discussed later) that guanine nucleotides-such as hydrolyzable analogues [Gpp(NH)p lated the affinity of this receptor quires

Am

J Psychiatry

1 49:6,

June

1992

cAMP)

proteins of exthe cell (3, 4). research of

adeny!y!

activity

antagonists. These and other indirect observations led to the proposal that the site of GTP’s action is on a protein distinct from both the hormone receptor and adenylyl cyclase. This hypothesis was confirmed by the subsequent purification of a GTP-binding protein from liver. By using the 549 cell mutant (deficient in hormoneand GTP-dependent activity), now termed “cyc,” it was possible to demonstrate that the addition of the purified “transducer protein” restores hormoneand GTP-dependent adenylyl

cyclase

activity.

Since

the

addition

of

this

pu-

rified protein stimulated adenylyl cyclase activity, it was termed “G,” (s=stimulatory) (reference 7 is an excellent historical overview). Shortly thereafter, the purification of a protein necessary for inhibition of adenylyl cyclase (“G1”) provided a mechanistic explanation for the bidirectional regulation of this enzymatic activity (8). G-like proteins were shown to activate a phospholipase C that hydrolyzes phosphatidylinositol 4,5bisphosphate to generate diacylglycerol and inositol 1,4,S-trisphosphate and to modulate the activity of specific

ion

channels

(3, 4, 9, 10).

Subsequently,

another

G

protein for which no obvious function was evident was purified from brain (11); this protein was named “G0” (o=other protein). The identification of G0 led to the realization that the G proteins are, in fact, a family of homologous proteins serving diverse roles in a wide variety of receptor-mediated extracellular signals to vanous intracellular second messenger systems (3, 4, 9, 10). It is now clear that G proteins involved in signal transduction are cx1y hetenotrimers. The greatest diversity observed thus far is with a subunits, which are generally believed to directly modulate the activities of vanous effectors (discussed later), but diversity also exists for and y subunits (see table 1).

MECHANISM

OF G PROTEIN

ACTIVATION

G

proteins in neurotransmitter-neurotransmitter interactions, and the possible involvement of G proteins in a variety of clinical conditions, with an emphasis on psychiatric conditions and their treatments.

WHAT

K. MANJI

re-

It was subsequently agonists stimulated in erythrocytes and GTP or similar nonor GTPyS]-modufor agonists but not

The molecular events underlying G protein activation/deactivation have been best characterized for Gprotein-mediated regulation of adenylyl cyclase (3, 4, 13), but the following sequence of events is assumed to occur for most (if not all) G proteins (see figure 1). The G protein a subunit cycles between an inactive GDP-bound oligomeric (4y) form and an active GTPbound monomeric form. Gilman (4, 13) has described these two forms of the a subunit as the “off” and “on” positions of a carefully timed molecular switch. Activation of a receptor by an agonist induces a conformational change in the receptor, allowing it to interact with the G protein, and forms a short-lived “high-affinity ternary complex” consisting of these components. The interaction of the receptor with the G protein in turn results in a conformational change in the a subunit of the G protein, which facilitates the displacement of GDP by GTP. The binding of GTP is a crucial step in the activation cycle and has two major consequences. 1 The G protein dissociates from the receptor. This destabilization of the high-affinity ternary complex re.

747

G PROTEINS

TABLE

1. Key Features

of G Protein Subunitsa

Subunit

Molecular Weight (x103)

Ga, (four types)

44.5-46

G, olf Ga1 (three

40.4-40.5

Toxin Targeting Subunit (cholera or pertussis)

Effector(s)’

Examples

Cholera

AC (+); L-type

Cholera

AC

(+)

Pertussis

AC

(-);

40.5

Both

Cyclic

Ga Ga0 (two types)

40.5 39.9

Both Pertussis

Cyclic GMP phosphodiesterase K channels (+); Ca2 channels PLC (+) (sensitive to pertussis

Gct

40.9

Neither

PLC

42

Neither

PLC (+) (insensitive

Direct interaction with AC (-)‘; inactivates a y required for interaction of a subunit with receptor

Ga

r

G

l

44.7

types)

,

G14

,

?four

types)

37.4

Neither

#{149}1 (three

types)

8-10

Neither

aAdapted from Birnbaumer (3), Taylor bAC=adenylyl cyclase, PLC=phospholipase CFinding uncertain.

et al. (9), Freissmuth C, +=stimulatory

sults

to the

in the

receptor’s

reversion

K channels GMP

low-affinity

con-

of

is terminated to

GDP.

the

However,

additional

adenylyl

ct-GDP

by the action

a subunit, this process amplification

of a GTPase

enzyme

which hydrolyzes GTP to is relatively slow, allowing of the signal. The forma-

is presumed

to

cause

dissociation

of

a

effector (which is now free to interact with activated a-GTP subunits); the reassociation with y is thermodynamically stable and the cycle with the formation of the inactive holo-G protein. appear to exert their inhibitory effects on cyclase

by two

distinct

mechanisms.

First,

in a

manner closely paralleling that just outlined for there appears to be a direct inhibitory effect of aj on catalytic unit of adenylyl cyclase. Gilman (4, 13) proposed that, in addition to the direct inhibition by the ‘y subunits, released by receptor activation of may by mass action attenuate the dissociation of Since the quantity of G is considerably greater than quantity of G, in most tissues and y subunits appear be largely interchangeable, this sequestration model received

much

acceptance.

Although

some

debate

G, the has a, G, G. the to has

per-

sists as to whether the a, or y subunits have the greater role in the inhibition of adenylyl cyclase, more studies suggest that they exert distinct but complementary effects

subunits

748

in

and Simon -=inhibitory

Unknown Thromboxane

A2, vasopressin

(12). effect.

stimulation

of

that

a is the primary

adenylyl

cyclase

(14).

mediator

It is

of inhibi-

active.

intrinsic

of

toxin) toxin) toxin)

tion of adenylyl cyclase when G is stimulated by a hormone or neunotransmitter but that y exerts a major role in the inhibition of basal and postreceptor-stimulated adenylyl cyclase activity.

a-GTP

from the additional of a-GDP completes GDP-bound Receptors

(-);

to pertussis

possible

The ticular, lected

tion

(+) (+)

to pertussis

dependent thus

12, D1, A2, H2, ACTH, CRH, V2, PGE1 Olfactory signals 2’ D2, A1, t, M2, S-HTIA Retinal rods (rhodopsins) Retinal cones (rhodopsins) a2, GABAB ,

(+)

(insensitive

(+)C

(+)

phosphodiesterase

2. The second major consequence of GTP binding to the G protein is to promote dissociation into a-GTP and ysubunits. It is generally accepted that, for a numben of effector systems, the a-GTP complex activates the effector enzyme by an as yet undetermined mechanism. The continued activation of adenylyl cyclase by

for

channels

et al. (10), and Strathmann effect of subunit on effector,

formation and the dissociation of the agonist from the receptor. Thus, the receptor is not permanently associated with the G-adenylyl cyclase complex, and this a!lows the receptor to recycle and function catalytically, activating several G proteins during the time that one

G remains

Ca2

of Receptors

the

may

inhibition

be more

of

potent

adenylyl

inhibitors

cyclase.

Thus,

of receptor-in-

‘y

G PROTEINS

AS TARGETS

discovery pertussis G proteins

various

subunits

TOXINS

that certain bacterial toxins (in parand cholera) can cova!ent!y modify sehas been invaluable in the delineation

structural

homologous and pertussis diphosphate

FOR BACTERIAL

and

functional

aspects

of

highly

G proteins. In brief, both cholera toxin toxin catalyze the transfer of an adenosine ribose (ADP-nibose) moiety onto the a

of various

G proteins.

These

ADP-nibosylation

reactions, and their subsequent effects on signal transduction, are believed to be responsible (at least in part) for the pathophysiology of the diarrhea and whooping cough caused by cholera toxin and pertussis toxin, respectively. A major difference between the stimu!atory

(Ga) and other

subtypes

of G proteins

is their

suscepti-

bility

to toxin-catalyzed ribosylation. The a subunits of G5 and G (transducin, the major retinal G protein) are substrates for ADP-nibosylation by cholera toxin (3, 13). G is not generally a target of cholera toxin; rather, it is preferentially ADP-ribosy!ated in an analogous manner by pertussis toxin. ADP-ribosy!ation by pertussis toxin appears to stabilize the G protein in its inactive ay undissociated conformation. Additionally, pertussis

toxin

thereby

“uncouples” attenuating

receptors

from

receptor-mediated

G1 (3,

7),

4,

inhibition

of

adenylyl cyclase. Studies shown that several G proteins sis toxin, namely, G1, G12, additional techniques have tween the various pertussis

to be used to distinguish betoxin substrates. Neverthe-

less,

of these

the

relative

selectivity

Am

using cDNA cloning have are substrates for pertus-

G3,

G,

J Psychiatry

and G0 (15).

toxins

I 49:6,

has

Thus,

resulted

June

1992

HUSSEINI

in their extensive use in identifying the presence and G, in various tissues, as well as the coupling ous receptors to these G proteins.

of G of van-

FIGURE

1. Mechanisms

K. MANJI

of G Protein Activation/Deactivationa Aonlst

High-Affinity

Ternary

Beta

Complex

Receptor

G PROTEIN

REGULATION

OF PHOSPHOLIPASE

The phosphoinositide pathway is a major signaltransducing pathway that has a ubiquitous role in the regulation of various aspects of cellular and neuronal function. The binding of hormones and neurotransmittens to a variety of cell surface receptors-e.g., a1-adrenergic, serotonin (5-HT1, 5-HT2), and muscaninic (M1, M3)-activates, through interposed G proteins, a phosphoinositide-specific phospholipase C (16). This enzyme hydrolyzes phosphatidyl inositol 4,5-bisphosphate (PIP2) to generate two second messengers: inosito! 1,4,5-trisphosphate (1P3), which mobilizes Ca2 from intracellular stores, and 1,2-diacylglycerol, which activates protein kinase C (16). Protein kinase C occupies a pivotal position in the biochemical pathways that relay information into the cell, because it is able to influence the cellular response to numerous other stimuli. Indeed, activation of protein kinase C has been linked to the

regulation

of cell

surface

receptors,

ion

elusive

G

proteins

coupled

to

stimulation

J Psychiatry

1 49:6,

June

1992

Subunit DissOciatiOn

IL__i

Subunit Reassociation

I

Affinity)

(Low

P1 GTPase

Intrinsic

-

GDP

Inactive

GTP Active

as

as

Adenyiate Cyclase

AlP

cAMP

M+

aAt

rest

an equilibrium

exists

between

the

receptor

in the

high-affinity

state (coupled to the G protein) and the low-affinity state. Activation of receptors by an agonist induces a conformational change in the receptor, allowing it to interact with the G protein, and forms a short-lived “high-affinity ternary complex” consisting of these components. The receptor-G protein interaction facilitates the replacement of GDP by GTP on the guanine nucleotide site (on the a subunit of the G protein). Binding of guanine nucleotides to the G protein causes a destabilization of the high-affinity complex and a dissociation of the G protein into a-GTP and 1y subunits. For most effector systems, the a-GTP complex activates the enzyme or ion channel. The continued activation is terminated by the action of a GTPase enzyme intrinsic to the a subunit, which hydrolyzes GTP to

GDP; the

the reassociation

formation

of the

cumulated pase

C,

that

of GDP-a inactive

similar

G proteins

with 1y completes

the cycle

with

G protein.

to the may

regulation

couple

various

of phospholireceptors

to

phosphatidylcholine by means of phospholipase (21 ). The existence of this signal transduction pathway has generated considerable interest, since it provides mechanistic explanation for the frequently reported temporal dissociation between production of diacylglyerol and 1P3 (21).

D a

of

phospholipase C in a pertussis-insensitive manner (12). Similar to the dual regulation of adenylyl cyclase, a number of receptors appear to inhibit phosphoinositide turnover, including receptors for adenosine, dopamine, S-HT, and glutamate (20). In addition, evidence has ac-

Am

GDP

Stimulatory G Protein Complex

channels,

secretion, gene expression, and neuronal plasticity (17). The details of the phosphoinositide turnover pathway have been outlined previously in an excellent review in the Journal ( 1 7) and will not be repeated here. The G protein that stimulates phosphoinositide turnover has been termed “Ge” (p=phosphoinositide), a!though its exact nature remains to be determined. What is clear, however, is that there appears to be more than one type of G protein activating phospholipase C, since pertussis toxin (which ADP-ribosylates and inactivates several G proteins [see preceding discussion]) inhibits agonist-stimulated PIP2 hydrolysis in some cell types but not others (16). Indeed, functionally distinct G proteins may even selectively couple different receptors to PIP2 hydrolysis in the same cell (18). Studies using Xenopus oocytes (large frog eggs frequently used in transfection studies) suggest that G0 rather than G may represent the pertussis-toxin-sensitive “Ge,” at least in certain cell types (16). The identification of a novel G protein termed “G7,” which is not a substrate for nibosylation by either cholera toxin or pertussis toxin, has led to the proposal that G may represent the “G” in a variety of tissues, including brain (where it is abundant) and platelets (19). More recently, however, several groups have purified and cloned members of a novel, ubiquitously distributed family of G proteins known as Gq. The a subunits of this family, including aq and , have unequivocally been shown to stimulate phospholipase C, making them leading candidates for the heretofore

(Low Affinity)

(High Affinity)

C

G PROTEIN

COUPLING

OF RECEPTORS

TO

ION

CHANNELS It has become increasingly large number of ion channels

clear in recent are regulated

years that a both directly

749

G PROTEINS

and indirectly by G proteins; at least 12 separate ion channels appear to be directly regulated (under cell free conditions) by G proteins (3, 22). In the CNS, a variety of receptors regulate neuronal K conductances by means of pertussis-toxin-sensitive G proteins. Activation of these K channels results in neuronal hyperpolanization and is an important element in the regulation of the neuronal firing rate (23). Although both a, and

a0 are able

to activate

inwardly

rectifying

K

currents

in neurons, a0 is markedly more potent. This is of considerable interest since a0 can be said to be predominantly localized in the nervous system (24), where its major role may be to regulate ion channels. Indeed, the most abundant G protein in the brain is G0, which cornpnises 0.S%-1.0% of the total membrane protein (24). Within the brain, G0 has a heterogenous distribution; it is particularly rich in the hippocampus and frontal contex but is also widely distributed in other areas of the brain (25). This distribution parallels that of protein kinase C (but not adenylyl cyclase) in the brain, leading to the suggestion that G0 may represent the pertussistoxin-sensitive “G” in the brain (25). In addition to regulating K channels, G proteins (in particular, G0) also directly modulate neuronal Ca2 channels (3, 22, 26, 27). Voltage-activated Ca2 channels are present throughout neurons and regulate a variety of cellular functions, including, of course, neurotransmitter release. A variety of neurotransmitter receptors (including a2-noradrenergic, GABAB, and adenosine Al ) inhibit neuronal voltage-sensitive Ca2 channels, and this receptor-stimulated modulation of Ca2 ion channels may be (at least in part) the molecular mechanism whereby nerve terminal autoreceptors inhibit neurotransmitter release (26, 27).

crease in Ga,, mRNA, suggesting in vivo physiologic regulation (33). It has also been demonstrated that pretreatment of mouse stniatal neurons in primary culture with 1 7 3-estradiol or testosterone increases the pertussis-toxin-catalyzed ADP-nibosylation of Ga0 and/on

Ga1 subunits

as evidenced

tion, and methasone

by

OF

G PROTEINS

BY

However,

it has

been

HORMONES

demonstrated

that

7-day

administration of corticosterone increases the transcniption and expression of Ga5 while decreasing both the mRNA and immunoreactivity of Gaj in rat cerebral cortex (33). Moreover, adrenalectomy without corticosterone replacement results in a significant 20% de-

750

ACTH

hypersecre-

of plasma cortisol after dexa(35). Similarly, at least some studies have shown the administration of thyroid hormones to be beneficial in the treatment of refractory depression and rapid-cycling bipolar disorder. Although a variety of biochemical effects may contribute to the CNS manifestations of abnormalities in thyroid or conticosteroid status, alterations of G protein function or content (with the inherent amplification of receptor responses) by these permissive hormones are an attractive mechanism. Similarly, the effects of gonadal hormones on G proteins, resulting in subsequent modification of signal transduction (34, 36), may be one mechanism by which biological maturation (puberty or menopause) triggers the expression of certain psychotic illnesses

cholic

(e.g.,

schizophrenia

depression

in late

in late

adolescence,

melan-

adulthood).

AS MEDIATORS

OF

NEUROTRANSMITfER-NEUROTRANSMITfER AND RECEPTOR-RECEPTOR INTERACTIONS

The CNS is a major target for the actions of glucocorticoids, thyroid hormones, and gonadal hormones, but the biochemical alterations ultimately responsible for producing the effects remain unclear. Malbon et a!. (28, 29) coined the term “permissive hormones” to describe agents such as glucocorticoids and thyroid hormones that modulate the actions of a variety of agents acting through cAMP. Increasing evidence suggests that these modulatory actions are exerted to a significant degree through G protein regulation. Thus, in vivo alterations of hormonal levels have been demonstrated to alter the steady-state levels of several G protein subunits and thereby regulate the overall sensitivity of transmembrane signaling in a variety of peripheral tissues (3032). Less information is available about the effects of hormonal manipulation on G protein regulation in the CNS.

hypercortisolemia,

nonsuppression administration

G PROTEINS REGULATION

(34).

Abundant evidence supports an interaction between thyroid and adrenal hormones and psychiatric illness, particularly in affective disorders. Thus, primary disordens of both the thyroid and hypothalamic-pituitaryadrenal (HPA) axis have been linked with depressive, manic, and anxiety symptoms. Additionally, there is general agreement that more than 50% of patients with major depression exhibit hyperactivity of the HPA axis,

ple

Most clinical measures

terms

studies in psychiatry, are obtained, analyze

of independent

measures,

e.g.,

even data

when multiprimarily in

Is norepinephnine

deficient? Is serotonin deficient? However, considerable preclinical evidence shows that monoamine systems interact, and a major question for neuroscience is emenging with regard to elucidating the mechanism(s) by which one neurotransmitten influences the response of a neuron to all the other converging afferent inputs. The CNS is extremely complex, both anatomically and chemically, and there is a remarkable convergence of different receptors in common cortical layers (37) and a considerable convergence of neurotransmitter action (38). A single neuron in the brain receives thousands of synaptic inputs on the cell body and dendrites, and neuronal response is also modulated by a variety of hormonal and neurohormonal substances not dependent on synaptic organization. The neuron needs to integrate all the synaptic and nonsynaptic inputs impinging on it; this integration of a multitude of signals determines the ultimate excitability, firing pattern, and response characteristics of the neuron, and the neuron’s

Am

J

Psychiatry

1 49:6,

June

1992

HUSSEINI

response is then conveyed to succeeding means of synaptic transmission. How does neuron decipher and integrate the multitude it receives and, additionally, generate unique to each

of these

signals

or combinations

of signals?

do G proteins

form work

the basis of a complex information processing netin the plasma membrane (39). Thus, the ability of

to interact

an elegant

signals,

with

mechanism

they

multiple

also

appear

receptors

to organize

the

signals

from

messenger

. NE

according

to

their

G protein

and

inte-

pathway.

Similarly, the dual (positive and negative) regulation of adenylyl cyclase and perhaps of phospholipase C by G proteins allows stimulatony and inhibitory signals for these pathways to be balanced at the G protein level, yielding an integrated, dampened output (39). Convergence

of a variety

of neurotransmitters

on

the

same

Alpha-2 Blockade

A,

and

and

same

the

hippocampus,

K-opiate pool

through creasing

receptors

of

G

is mutual

appear

proteins

voltage-sensitive norepinephnine

there

a2-adrenergic, to

modulate

Ca2 release

antagonism

Ca2

channels, (44-46).

for

these

de-

2).

Similarly,

heterologous

of desensitization

agent leads of stimuli, nisms) may protect the stimulation provide the receptors to tions linking with

signals

exposure

to

(a form a desensitizing

to diminished responsiveness to a number apparently through postreceptor mechabe a compensatory mechanism designed to neuron from the deleterious effects of overby multiple receptors. Thus, G proteins first opportunity for signals from different be integrated; this complex web of interacreceptors, G proteins, and their effectons converging to shared detectors appears to

be crucial

for the

CNS

47,

(39,

desensitization

whereby

integrative

functions

Kappa opiate

receptors

with respect to modulating both Ca2 influx and nonepinephnine release. That is, stimulating one of these receptors diminishes the effectiveness of the other receptons; this may be a mechanism for allowing neurons to “escape” from excessive inhibitory input (46) (see figure

NE

the

influx

thereby Additionally,

between

Alpha-2 Stimulation

adenosine

to compete

NE

ion

channel has been demonstrated to occur in the locus coeruleus, hippocampus, thalamus, and substantia nigra (38, 40-43). Thus, it appears that a variety of receptons which coexist on neurons and mediate similar responses may share signal transduction mechanisms (38, 40, 43, 44). This convergence may occur at the level of the G protein or the effector itself. In both the cortex

performed

by the

48).

aReceptors

that

coexist

EFFECTOR

CROSS-TALK

In addition

G proteins between

Am

also various

J Psychiatry

OF RECEPTOR-

IN THE

to their role as receptor-effector serve as targets for regulating second messenger systems.

I 49:6,

June

1992

mediate

number of potential sites of interaction ase C with the cAMP-generating system ure

3).

rylation

A large of the

body

of evidence

3-adrenergic

similar

responses

biochemical

effect of -adrenergic

by cross-talk evidence

that

phospho-

by various

kinases,

desensitizes the receptor that the most commonly of antidepressants, receptors,

through

protein

has

accumulated

also

of protein kin(49, 50; see fig-

suggests

receptor

including protein kinase C, (50). Indeed, evidence suggests

Increasing

couplers, cross-talk There are a

and

possibly by making available a greater proportion of the G protein pool. In an analogous manner, stimulation of a2 (bottom) diminishes the effectiveness ofthe other receptors a disproportionate use of the common pool of G proteins. This may be a mechanism for allowing a neuron to “escape” from excessive inhibitory input.

diated

CNS

neurons

(middle), shared receptors through

down-regulation

AS MEDIATORS

on

may share signal transduction mechanisms. The top portion of the figure depicts a neuron with a,-adrenergic, adenosine A,, and K-opiate receptors all converging onto a common pool of G-like proteins to inhibit Ca2 influx and norepinephrine (NE) release. Alpha-2-adrenergic antagonism increases the effectiveness of the other receptors

observed

G PROTEINS

by a Corn-

these

grated

second

Interactions

provides

them to a relatively Signals from a van-

a single

of Receptor-Receptor Systerna

to

multiple receptors and to transmit much smaller number of effectors. ety of receptors can be “weighted” intrinsic ability to activate a given to stimulate

FIGURE 2. Integration mon G Protein-Effector

Not

only

G proteins

amplify

targets by the single of signals responses

K. MANJI

kinase

namely, may

be me-

C (51, that

the

52). po-

tentiating effects of protein kinase C on cAMP accumulation are mediated by G. Thus, in a number of cell types, including hepatocytes and stniatal membranes (30, 53-55), activation of protein kinase C appears to

751

G PROTEINS

FIGURE 3. Mediation G Proteinsa

of Receptor-Effector

Cross-Talk

in the CNS by

duction pathways, it is not surprising that abnormalities in the function and/or expression of various G proteins have been implicated in a variety of pathophysio!ogic states: pseudohypoparathynoidism type I, heart failure, certain endocrine tumors, McCune-Albright syndrome, diabetes, alcoholism, schizophrenia, mitral valve prolapse, chronic cocaine/opiate ingestion, aging, hypo/hyperthyroidism, and adrena!ectomy/corticosteroid administration (58-81). It should be noted that G protein dysfunction appears to be the primary pathology in Albright’s hereditary osteodystrophy, McCune-Albnight syndrome, and endocrine tumors; in several of the other conditions, G protein abnormalities are probably secondary but are nonetheless implicated in the pathophysiology of the condition. I will briefly highlight some of the clinical conditions with well-characterized

G protein dysfunction. The first disease in which Calcium

normality was called because

Mobilization

ma! (or even aThe

figure

depicts

ceptors

activating

ceptors

coupled

pholipase

Ca2

the

C results protein

interaction

intracellular

the phosphoinositide

to adenylyl in the

from intracellular

tivates

complex

kinase

of protein

cyclase. generation

stores, C. There

kinase

Protein kinase an uncoupling rylate adenylyl

C phosphorylates of the receptor

on differences

among

cross-talk

turnover

Receptor

of IP3 (IP3),

are a number

of protein

kinase

subtypes

which

(DAG),

of potential

re-

and re-

of phos-

sites

ac-

for the

system.

the 3-adrenergic receptor, causing from G,. It also appears to phospho-

of G proteins

of adenylyl cycit appears to phosits inhibitory tone. probably depends

and the relative

abun-

C isozymes.

enhance adenyly! cyclase activity by attenuating the inhibitory influence of G. Since the susceptibility of G proteins to be phosphorylated (and therefore regulated) by protein kinase C depends in large part on their conformational state (56), the degree of cross-talk is negulated by the relative degree of simultaneous stimulation of various receptors. In sum, it is clear that interactions between distinct second-messenger-generating systems represent a fine-tuning cellular network that regulates the neuron’s reactions to the large number of extracellular signals it encounters. The net effect of the various potential interactions probably depends on the summation of the effects on individual components. In this context, quantitative and qualitative (e.g., conformation states) differences among subtypes of G proteins and the relative abundance of protein kinase C isozymes (57) in various cells may be major factors in determining the final integrated output.

CLINICAL

752

their widespread regulation, and

and crucial amplification

role in the inteof signal trans-

ab-

have

so nor-

than

normal)

levels

of parathyroid

demonstrate

a resistance

to parathynoid

hormone and to a number of hormones using cAMP as the second messenger. In these patients, lower than norma! expression or function (approximately 50%) of Gcx, has been demonstrated in cultured cells and in cells from freshly obtained tissue (58 and references therein). Additionally, distinct mutations have been identified in the gene encoding Ga in different kindreds with the disease (58, 59), thereby providing the first demonstrations of an inherited mutation in a human G protein gene with important implications. The existence of more than one type of mutation resulting in abnormal function of Gas, demonstrates the genetic heterogeneity of this autosoma! dominant disease. Moreover, the presence of an identical mutation in the Ga,, gene, both in individuals with multiple hormone resistance and in those without hormone resistance, clearly suggests that

G5 deficiency

is necessary

but not sufficient

for the full

phenotypic expression of the disease (59). Thus, as has been suggested for a variety of psychiatric disorders, additional factors, including modifying genes and perhaps environmental factors, appear to be involved. Studies have also demonstrated substantial abnormalities in G proteins in failing human and animal heart. A 50% lower than normal apparent concentnation and function of G in sarcolemma from a canine model of left ventricular failure has been reported. In contrast, end-stage idiopathic human congestive failure is associated with greater than normal activity of G, suggesting that various manipulations of G/G1 stoichiometry may result in similar pathophysiology (60). Perhaps more akin to psychiatric research with respect cessible

Given gration,

higher

to the use of peripheral

IMPLICATIONS

G protein

hormone but have the manifestations of the disease because of cellular resistance to the hormone. In one form of the illness (Albright’s hereditary osteodystrophy),

the patients

mobilizes

which

C with the cAMP-generating

cyclase, resulting in an enhancement lase activity. Finally, at least in certain systems, phorylate and inactivate G, thereby removing The net effect of the various potential interactions dance

pathway

activation

and diacylglycerol

between

an intrinsic

found is pseudohypoparathyroidism, individuals with this condition

80% jects

tissue

lower with

blood

(CNS,

than congestive

cells to represent

heart),

normal

study

lymphocyte

heart

Am

one

failure.

J

Psychiatry

(60)

less acshowed

G5 levels More

intriguing,

1 49:6,

June

in subsuc-

1992

HUSSEINI

cessful treatment with captopnil was associated significant two-fold increase in lymphocyte G

The following ciation

are summaries

of G proteins

of research

to psychiatric

with levels.

a

on the asso-

conditions.

The evidence for genetic factors in alcoholism is compelling (62), and, given the widespread medical, behavioral, and societal manifestations of this disorder, it is not surprising that there has been considerable research

factors

that may predispose

individuals

to

alcoholism. A variety of putative electrophysiologic, biochemical, and neuroendocrine markers have been studied, but increasing research has focused on G proteins not only as the mediators of the biochemical effects of alcohol but also as possible sites of underlying pathophysiology in individuals predisposed to the development of alcoholism (63). Ethanol can profoundly affect the G protein-adenylyl cyclase signal transduction pathway (63). Chronic ethanol ingestion or in vitro exposure results in a desensitization of cAMP production in a variety of tissues, including brain (63, 64). The heterologous nature of desensitization of the adenylyl cyclase activity following chronic in vitro exposure of neuroblastoma cells to ethanol suggests a postreceptor (e.g., G,) site. Chronic exposure of cultured cell lines to ethanol results in a more than 30% decrease in both the a protein and its mRNA (65, 66). In mice chronically fed ethanol, there is a reduction in the cholera-toxin-catalyzed [32P]ADPribosylation of G and a reduction of high-affinity [3H]forskolin binding (which is thought to reflect binding to the complex of a and adenylyl cyclase) (63). Clinical studies have also implicated G-adenylyl cyclase as a putative marker in alcoholism. One study showed that basal and adenosine-receptor-stimulated cAMP 1evels

in fresh

lymphocytes

of alcoholic

subjects

were

75%

lower than those of normal subjects or subjects with nonalcoholic liver disease (67). Cultured lymphocytes from alcoholics, grown in the absence of alcohol, also show some abnormalities in the cAMP signal transduction pathway (64). Similarly, platelet membranes of alcoholics reportedly abstinent from alcohol for 12-48 months demonstrate lower receptorand G5-stimulated adenylyl cyclase activity than do platelet membranes of age- and sex-matched comparison subjects (68). These findings suggest that low platelet adenylyl cyclase activity may be

a genetic by studies

marker by my

in alcoholics; colleagues

tussis-toxin-catalyzed

this finding and

me

of cholera-

[32P]ADP-nibosylation

G, and G, respectively,

in platelet

is supported and

per-

of platelet

membranes

from

ab-

stinent alcoholics, subjects with concomitant diagnoses of major depression and alcoholism (major depression as the primary diagnosis), and age- and sex-matched companison subjects. We observed significantly less 32P-labeling of G (but not G#{149}) in the alcoholic group than in the comparison subjects. These findings are entirely consistent

with

koff

et a!. already

Am

the

J Psychiatry

platelet

adenylyl

described

1 49:6,

cyclase

(68).

June

1992

findings

Opiate/Cocaine

To my knowledge,

Use there have been no clinical

studies

of the gestion clinical

Alcoholism

on biological

Chronic

K. MANJI

by Taba-

effects of acute or chronic opiate or cocaine inon G proteins. Nevertheless, considerable preevidence suggests that these substances exert at least some of their chronic effects (e.g., tolerance, dependence, withdrawal) through G proteins. A variety of opiate receptors are coupled to their effectors by G proteins, and pertussis toxin uncouples opiate receptors from the inhibition of adenylyl cyclase in cultured cells and in locus coeruleus neurons (3). Studies have shown that alterations in G protein levels are temporally conrelated with neurona! activity in the locus coeruleus (more firing) and with the behavioral manifestations of the morphine withdrawal syndrome in rats (69, 70). These results suggest that alterations in the levels of G proteins may underlie the enhancement of neuronal excitability (and possibly withdrawal symptoms) observed after abrupt opiate discontinuation. In vitro studies of cultured neurons from rat spinal cord dorsal root ganglia have demonstrated that exposure to K-opiate agonists is accompanied by a 60%-70% decrease in levels of aj, in the absence of alterations in levels of a,

a0, and

subunits

(71),

thereby

providing

tic explanation for opiate-induced sitization. In contrast to the studies recent study has directly examined

G proteins, ventral coeruleus

demonstrating

tegmental in rat

low levels

a mechanis-

heterologous on opiates, cocaine’s

desenonly one effects on

of a and a0 in the

area, nucleus accumbens, brain after chronic cocaine

and locus use (72).

Schizophrenia Several years ago, studies brain tissue demonstrated ylyl

cyclase

activity

using post-mortem a greaten response

to fluoride

stimulation

human of aden(presumably

acting through G) in the caudate and accumbens of schizophrenic patients than in the corresponding brain areas of comparison subjects (73). These studies suggested that a postreceptor defect may contribute to the presumed dopaminergic hyperactivity in schizophrenia, but it was several more years before they received additional, more direct support. Seeman et al. (74) used radioligand binding studies to explore the D1-D2 link in post-mortem human brain tissues. They proposed that the D1 receptor may modulate ligand binding to the D2 receptor by modifying the levels of G protein subunits

(1)

that these

receptors

share

(see figure

4), and they

reported

that the D1-D2 link was missing in more than 50% of tissue samples from patients with schizophrenia and Huntington’s disease, a finding apparently not due to prior neuroleptic use. In a study that examined G protein levels in post-mortem schizophrenic brains more directly, pertussis-toxin-catalyzed nibosylation in the left putamen was 42% lower in schizophrenic patients than in comparison subjects (75). These findings (suggesting low G1 and/or G0) would, of course, be compatible with enhanced dopamine-adenylyl cyc!ase function. In keeping with these lateralized abnormali-

753

G PROTEINS

FIGURE

4. Reciprocal

Modulation of Receptor Affinity by G Proteinsa

ribosylation

of GIG0

sent

1MB

link

D1-D2

Two

studies

jects

GDP

inactiveai

as

cussion). strated

02 AgonIst

U

in depressed

84),

abnormalities

ing

these

allows

further

interac-

released may drive the equilibrium for another G protoward the undissociated state and thereby modulate receptor affinity (in this case, the D, receptor). This may be one mechanism for the observed synergism between the D, and D2 receptors despite opposite effects on adenylyl cyclase. (e.g.,

thus G)

For

ties

in postreceptor

pathways

is thought adenylyl

to

reflect

cyclase,

in schizophrenic

binding higher

than

patients,

[3H]forskolin bindparahippocampal gyrus brains of schizophrenic [3H]forskolin binding to

the normal

complex levels

of of

a and a, and!

or adenylyl cyclase would be the simplest explanation for these results. However, low levels of G1 3y subunits would also result in a greater availability of free

a

to interact

with

adenylyl

cyclase.

Indeed,

although

this suggestion is highly speculative, a low level of y subunits would be compatible with the higher than norma! adenylyl cyclase activity (73), lower pertussis toxin

754

platelet

manic-

[3HJ-

are

to

in reference interpret

cAMP

levels)

lower

lymphocyte

accumulation

(85).

demonstrated

and

in in dewas

Similarly,

a significant

correlations

between

release and basal cyclase activity

platelets

(unpublished

urinary and postin both

observations).

It is presently unclear whether these abnormalities receptor and postreceptor sensitivity are primary the sequelae of abnormalities in circulating levels echolamines cultured

and/or cells

needed

ob-

patients with high circulat-

terminal insomnia and blunted accumulation in depressed patients (86). Additionally, my colleagues

measures of norepinephnine receptor-stimulated adenylyl lymphocytes

given

documenting abnormalities and glucocorticoids

in a subgroup of depressed agitation (and presumably

I have

rereceptor

(reviewed difficult

significantly

between cAMP observed

demon-

a2-adrenergic

patients

example,

from

to examine

confounding

greater than normal high-affinity ing has been found in the left and CA 1 region in post-mortem patients (76 ). Since high-affinity

from

isoproterenol-stimulated

literature catecholamines

catecholamine

and

tion of G proteins. When receptors coexist on neurons, a potential mechanism for modulation of receptor affinity by other receptors exists. Binding of an agonist to a receptor (e.g., D1 receptor) promotes the dissociation of the G protein into a and ly subunits. The tein

and

sensitivity

correlation lymphocyte has been

of 3y subunits

tissue

a number of studies have in lymphocyte -adrenergic

sensitivity

served only psychomotor

D2 Receptor

13’)’units

brain

isoproterenol-stimulated

(Low affinity)

interchangeability

G proteins in afreported that the

significantly

by

Although abnormalities

pression. cii

or ab-

higher than those These findings are intriguing of Schreiber et al. (83), who G proteins in manic sub-

were

determined

the extensive circulating

‘p affuiity)

minimal

examined al. (82)

et

in post-mortem subjects

ceptor

(High

be fewer

GTPyS binding in leukocytes. Although these findings need to be replicated, they are exciting since lithium is reported to attenuate G protein function (see later dis-

)GDP

GD$

relative

directly

of comparison subjects. since they parallel those reported “hyperfunctional”

/

would

heterotnimers-the

and greater [3H]forskolin binding from schizophrenic subjects (76).

Young

of Ga

depressive

aThe

have

disorders.

levels

D1 Agonist

GTP#{231})

there

Disorders

fective

Active

(74), tissue

in post-mortem Affective

as

(since

subunits available to form ay substrate for the nibosylation reaction),

D1 Receptor

inactive

(73)

1-

stress these

hormones. patient

possible

effects

of

Thus,

populations

G protein circulating

defects

in or are of cat-

studies are

of

clearly

free

of the

catecholamines

and

hormones. Similarly, a common postreceptor abnormality (e.g., at the level of G proteins) would provide a mechanism for depressed patients’ blunted growth hormone and prolactin responses to a variety of provocative

challenges

(35),

but

such

an

abnormality

remains

to be established. Other

Neuropsychiatric

Conditions

Lower than normal levels of platelet basal and postreceptor-stimulated adenylyl cyclase activity have been reported in both panic disorder (87) and posttraumatic stress

disorder

possibility dary

(88).

that

to abnormalities

these

Once

again,

platelet in plasma

Am

J

one

is faced

abnormalities

with

are

catecholamines

Psychiatry

1 49:6,

the

seconand/or

June

1992

1-LUSSEINI

stress

hormones.

symptoms

Another

illness

in common

hyperadrenergic

with

with

panic

dysautonomia

a constellation

disorder

is a form

associated

with

of

duced

[3H]GTPyS

of

manic

patients

mitral

valve prolapse (77). Elegant reconstitution studies have recently demonstrated that upon reconstitution into cyc lymphoma cells, erythrocyte G, from patients with mitral valve prolapse is hypenresponsive with respect to both adenylyl cyclase activation and receptor coupling (78). If a similar G abnormality is present centrally in panic disorder, it would provide an attractive mechanistic explanation (i.e., desensitization of 3-adrenengicreceptor-coupled adenylyl cyclase) for the therapeutic efficacy of both tnicyclic antidepressants and monoamine oxidase inhibitors in this condition. Finally, abnormalities in G proteins have also been implicated

in

disease

(80),

Huntington’s

and

G PROTEINS

disease

aging

(79),

Alzheimer’s

(81).

AS TARGETS

OF PSYCHOTROPIC

DRUGS

Lithium Although

lithium

recurrent its

is widely

affective

disorders,

mood-stabilizing

ever,

system

depletion

suggests

and

the

(since

blood-brain

barrier,

phosphatase phoinositide

could deplete turnover) (91,

sive

research

on

cyclase. In rat vivo (i.e., after has

been

adenylyl pressants,

exerts sys-

of the

does

not

inhibition

of

inosito!

the

inositol-1-

inositol and reduce 92). This hypothesis

phoshas

shared

namely, reported

Am

J

effects

attenuate

on

neuronal

adenylyl

both in vitro ofthe animal), receptor-

Gpp(NH)p,

and

and ex lithium

postrecep-

fluoride,

(93-96). In contrast has been reported

by these

two

signal

forskolin]

to most to induce

transduction

In this context, attenuate the binding

and carbachol et a!. (83)

Psychiatry

lithium

delays

antidefew, if

in response

(97). observed

1 49:6,

June

Using greater

1 992

systems,

lithium has agonist-induced to

both

been in-

isopno-

a similar method, isoproterenol-in-

from

subjects

with

lithium.

activation

untreated

In renal

of

bi-

or euthymic

epithelial

adenylyl

cyclase

by

Gpp(NH)p, which is compatible with attenuation of the formation of the active Gpp(NH)p-cx subunit (98). Similanly, the inhibitory effects of chronic lithium use on rat brain adenylyl cyclase are reversed by increasing concentrations of GTP (93, 94). Taken together, these results

suggest

that

the

physiologically

relevant

effects

of

lithium on adenylyl cyclase may be exerted at the level of G proteins (presumably at a GTP-responsive step). My colleagues and I have recently demonstrated significant increases in basal and postreceptor stimulated adenyly! cyclase activity in platelets (but not lymphocytes) obtained from normal volunteers after 14 days of lithium administration (95, 96). Since inactivation of G appears to exert significant effects on adenylyl cyclase activity in platelets but not in lymphocytes (96), we postulated that the striking tissue-specific effects could be explained by a lithium-induced attenuation of G function. To examine this more directly, we used platelet membranes from these same subjects to quantitate G proteins by using specific lyzed [32P}ADP-nibosylation.

antibodies We did

and

toxin-cata-

not observe any abnormalities in immunolabeling of platelet G but did observe a significant 37% increase in pertussis-toxincatalyzed [32P]ADP-ribosylation of platelet G. Since the undissociated, inactive afry heterotnimeric form of gest

that

for pertussis

lithium

inactivates

toxin,

G

by

our results

stabilizing

the

sugmac-

tive, undissociated conformation. Corroborating our human platelet findings, we have recently observed similar increases in pentussis-toxin-catalyzed [32P]ADPnibosylation in rat cortex after 4 weeks of lithium use but no alterations in the immunolabeling of G, G1, or

G0. Similar

Whatever

in [3H]GTP

terenol Schneiber

cells,

lithium’s

G proteins. to markedly

crease

stabilized

question, however, since alterations in have not been consistently found after (see 92). There has also been exten-

any, changes in the density of rat brain -adrenergic receptors. Lithium also attenuates receptor-stimulated phosphoinositide turnover in rat brain and in human platelets (reviewed in references 92 and 96). Since lithium affects both phosphoinositide turnover and adenylyl cyclase, recent attention has focused on mechanisms

patients

G1 is the substrate

cross

in leukocytes

in normal

polar

laboratory mechanism rently under with Mg2

[GTP, cyclase lithium

for

How-

lithium

generation

lithium

to

ton-stimulated

that

inositol

brain preparations, chronic treatment

shown

basis

unknown.

inhibits the activity of the en(89, 90) has resulted in exthe phosphatidylinositol signal

hypothesis

been called into brain PIP2 levels lithium treatment

of

second-messenger-generating that lithium, at therapeutically

relevant concentrations, zyme inositol-1-phosphatase tensive research on transduction

treatment

biochemical

remains

evidence

effects on discovery

for the

the

actions

accumulating

substantial tems. The

used

binding than

K. MANJI

thus

findings

been

the mechanism,

multiple

cyclase,

have

reported

by another

(R.S. Jope, personal communication). The by which lithium may inactivate G is curinvestigation and may involve competition on cross-talk with protein kinase C (99). signal

by affecting

transduction

phosphoinositide

G proteins

mechanisms

turnover)

and

(adenylyl

common

to

many

different neurotransmitters, lithium may be in a unique position to affect the functional balance between neurotransmitter systems. One might speculate that it is precisely this effect on the functional balance between interacting neurotransmitter systems that underlies lithium’s mood-stabilizing effects. Antidepressants Despite nisms

of

extensive action

research,

of antidepressant

the

molecular drugs

mecha-

have

not

been

clearly established. The most commonly observed biochemical effect of most clinically effective antidepressant treatments is a reduction in the norepinephninestimulated production of cAMP (desensitization), usually

(but

not

invariably)

accompanied

by

a reduc-

755

G PROTEINS

tion

in

the

number

of

3-adnenengic

receptors

(down-

CONCLUSIONS

regulation) (100). This has generally been regarded as an adaptation to the elevation of intrasynaptic norepinephnine through pnesynaptic mechanisms (e.g., reuptake blockade or monoamine oxidase inhibition). However, a direct postsynaptic effect of these drugs has

been

vitro

suggested

glioma

of

cells

(102,

to

receptor

have

attempted

sants

at the

to cause

G

by the

exposure

103)

to level

a functional state

adenylyl

to tnicyclic

cyclase,

effects of the

interferes

perhaps

f

receptor

with

(103,

105).

from

the high-afactivation

decreasing

the

Okada

affinity

of

et a!. (106)

by pertussis

sensitizing

toxin

simply

masks

desipramine’s

de-

In studies using high in vitro jtM), it has been demonstrated

concentrations that a variety

(50-300

of antidepressants interfere with G activation of adenylyl cyclase (107). Interestingly, greater inhibition was observed when the antidepressants were added before, rather than after, the addition of GTP; this finding is also compatible with an antidepressant-induced inhibition of G protein dissociation. However, given the high doses of antidepressants used, the physiologic relevance of these findings remains unclean. It was reported (in an abstract) that chronic administration of imipramine decreased

the

levels

ADP-ribosylation, These results are of some (109-111) gesting

of Ga

immunolabeling,

cholera

toxin

and Ga, mRNA in rat brain (108). difficult to reconcile with the results but not all (104, 112) studies, sug-

increased

postreceptor-stimulated

adenylyl

cyclase activity in rat cortical membranes after administration of antidepressants. Nevertheless, it remains possible that antidepressants attenuate -adrenergicmediated activation of G, while enhancing the effects of agents operating by pathways independent of the adrenergic receptor. Like most antidepressants,

(ECS) cyclase

desensitizes activity

the in rat

electroconvulsive

f3-adrenergic-mediated cortex

and

shock

adenylyl

hippocampus

(100).

Fewer studies have investigated the effects of repeated ECS on postreceptor sites. However, repeated ECS is reported to decrease both the agonist-induced [3H]GTP (113) and [3H]forskolin (114) binding in rat cortex. In a study using [35S]GTPyS binding (115), repeated ECS was

found

hippocampus to increase

756

mechanisms

to decrease

binding

in prefrontal

cortex

but to have no effect in the stniatum I35SIGTP’yS binding in the amygdala.

and

and

specific

clearly

come

the

a long

way

family.

The

neuronal

in the

past

diversity

and

has

clear

decade.

In particu-

and

family

serve the critical role extracellularly genertransmitting

these

inte-

forming the basis for a network (39). of G protein in a given the influence of physipharmacologic

implications

both

of the G protein

then

pathophysiologic,

tions,

of a neurotransmit-

progress in defining of the signal-transducing

grated signals to effectors, thus complex information processing The finding that the amount tissue is not static, but is under ologic,

a

the

surface is translated into a a physiologic) effect has

The G proteins or attenuating

signals

plays

Understanding

binding

lan, there has been dramatic the structure and the function continues to grow. of first amplifying

receptors

activation.

by which

not

only

perturba-

for

research

into

the etiology of various psychiatric conditions but also for the development of better treatments. Abnormalities in G protein function have now been identified in the etiology/pathophysiology of a variety of medical diseases. With the exception of chronic alcohol exposure,

opiate

neuroleptic

effects.

through

in neuronal

ten to a receptor at the cell biochemical (and ultimately

ated of

reported that pertussis toxin treatment of rats overcomes desipramine-induced receptor desensitization (as assessed by measurements of isoproterenol-stimulated adenylyl cyclase activity) without attenuating (in fact, promoting) desipramine-induced receptor downregulation. While these results may suggest that G and/or G0 are involved in desipramine’s effects, it is equally plausible that the removal of the inhibitory

tone

transduction

role

G protein

appears

subsequent

by

nucleotides

leads

of antidepres-

uncoupling and

C6

investigators

Desipramine

breakdown

G for guanine

the

and

in

and

antidepressants

G proteins.

(104)

chronic

(101)

Other

examine of

that

fibroblasts

down-regulation.

in rat cortex

finity

observations

human

Signal

central

withdrawal,

and

exposure,

the

dysfunction in psychiatric rect. Nevertheless, given

of G proteins seems

likely

yield

insights

in the that

schizophrenia!

for

postreceptor

disorders is currently the widespread, critical

regulation

of neuronal

future,

more

the

involvement

into

perhaps

evidence

less

diroles

function,

sophisticated

it

studies

of G proteins

will in the

pathogenesis of various major psychiatric illnesses. These putative G protein abnormalities may be subtle but sufficient to modify G protein functioning in response to neurotransmitter changes, permissive hormones, or other environmental events or stressors. Finally, given the increasing evidence that the currently available psychotropic drugs affect G proteins, the development of novel drugs with primary G protein targets

remains

an exciting

prospect

for

the

future.

Indeed,

it is intriguing that a number of the currently pharmacologic agents, although not developed appear to modify physiologic events that are

G proteins.

These

include

hepanin

available as such, linked to

derivatives

(116),

volatile anesthetics ( 1 1 7), and caffeine-like agents (118). It has also been demonstrated that a variety of cationic amphiphilic neuropeptides activate G proteins in a receptor-independent manner (119); they represent a novel class of agents (receptomimetics, rather than receptor agonists). Interestingly, tricyclic antidepressants are also cationic amphiphilic compounds (95, 120), so they

too

tion

to

may allow

adopt for

a membrane-spanning an

interaction

with

conformathe

guanine

cleotide binding site on the G protein. Recent have also identified a novel class of agents that allostenic enhancers and nist-preferring conformation

stabilize the high-affinity agoof the adenosine A1 recep-

ton (121). This may be viewed manner in which benzodiazepines

Am

nu-

reports serve as

J

as being analogous to the potentiate GABA-er-

Psychiatry

1 49:6,

June

1992

HUSSEINI

gic

neurotransmission.

Such

agents

have

a built-in

safety mechanism, since the maximal effect of the drug depends on the availability of the endogenous ligand. This may be one mechanism underlying the relative safety of benzodiazepines in overdose. Additionally, the use of such allostenic modulators may serve to effectively re-equilibrate dysregulated neuronal systems (which

are

likely

without the tachyphylaxis sensitivity.

to be involved

commonly (receptor Finally,

in psychiatric

encountered down-regulation)

given

the

fact

that

a fixed niques stimulate

phenomena and

super-

GTPase

activ-

the

of

as a “clock,” allowing the system to cycle at rate, it seems likely that molecular biologic techwill lead to the development of agents that either or inhibit

GTPase

by altering

Baraban

18.

and psychoactive drug action: focus on the phosphoinositide system and lithium. AmJ Psychiatry 1989; 146:1251-1260 Ashkenazi A, Peralta EG, Winslow JW, Ramachandran J, Capon DJ: Functionally distinct G proteins selectively couple different receptors to P1 hydrolysis in the same cell. Cell 1989;

its catalytic

rate,

thereby dampening or amplifying the hormonal (without constitutively activating the G protein). Appendix 1 provides more specific information methods for G protein research.

signal

19.

56:487-493 Casey PJ, Fong cleotide-binding

20. 21.

Exton

22. 23. 24.

25.

2131 I 1 . Birnbaumer ofeffector

1989;

13. 14.

15.

16.

Am

Psychiatry

June

1992

Signaling

Brown

AM,

Birnbaumer

by G protein

subunits.

that

inhibit

Sci 1989; phosphatidylcholine

through

1990;

340:

a guanine properties.

nu-

J

phosphoinosi10:114-120 breakdown.

265:1-4 L: Ionic

Annu

channels

Rev Physiol

and their regulation 1990; 52:197-213 and neuronal function. ofthe

Go protein.

PF, BarabanJM, Snyder SH: Beyond receptors: systems in brain. Ann Neurol 1 987;

Soc

multiple 21:217-

modulation of calcium currents in neu1990; 52:243-255 5, Tsien RW: Alpha-adrenergic inhineurotransmitter release mediated by calcium-channel gating. Nature 1989;

639-642

Ros M, Northup JK, Malbon CC: teins and beta-adrenergic receptors

29.

effects

ofthyroid

Malbon

CC,

hormones. Rapiejko

PJ,

30.

31

.

Steady-state levels of G-proin rat fat cells: permissive

J Biol Chem Watkins

regulation of hormone-sensitive macol Sci 1988; 9:33-36

1988;

DC:

effector

Houslay MD, Milligan G: G-Proteins Signalling Processes. New York, John Levine MA, Feldman AM, Robishaw

263:4362-4368

Permissive

hormone

systems.

Trends

Phar-

as Mediators of Cellular Wiley & Sons, 1990 JD, Ladenson PW, Ahn

TG, MoroneyJF, Smallwood PM: Influence ofthyroid hormone status on expression ofgenes encoding G protein subunits in the rat heart. J Biol Chem 1990; 265:3553-3560 32.

Ros

M,

Northup

adenylate

cyclase:

JK,

Malbon CC: Adipocyte of adrenalectomy.

G-proteins Biochem

effects

J

and I 989;

257:737-744

33.

34.

Saito N, Guitart X, Hayward M, Tallman JF, Duman RS, Nestier EJ: Corticosterone differentially regulates the expression of Gs alpha and Gi alpha messenger RNA and protein in rat cerebral cortex. Proc NatI Acad Sci USA 1989; 86:3906-3910 Maus M, Homburger V, Bockaert J, Glowinski J, Premont J: Pretreatment of mouse striatal neurons in primary culture with 17 beta-estradiol

ribosylation

enhances

ofG

alpha

the

pertussis

o,i protein

toxin-catalyzed

subunits.

ADP-

J Neurochem

1990;

SS:1244-12S1 35.

Nemeroff

36.

ogy. New York, Maus M, Bertrand

CB: Handbook

Glowinski

L: Transduction of receptor signal into modulation activity by G proteins: the first 20 years or so. FASEB

1 49:6,

JH:

Dolphin AC: G protein rons. Annu Rev Physiol Lipscombe D, Kongsamut bition of sympathetic modulation of N-type

28.

3:2125-

Strathmann M, Simon MI: G protein diversity: a distinct class of alpha subunits is present in vertebrates and invertebrates. Proc Natl Acad Sci USA 1990; 87:9113-9117 Gilman AG: The Albert Lasker Medical Awards: G proteins and regulation of adenylyl cyclase. JAMA 1989; 262:1819-1825 Hildebrandt JD, Kohnken RE: Hormone inhibition of adenylyl cyclase: differences in the mechanisms for inhibition by hormones and G protein beta gamma. J Biol Chem 1990; 265: 9825-9830 Jones DT, Reed RR: Molecular cloning of five GTP-binding protein cDNA species from rat olfactory neuroepithelium. J Biol Chem 1987; 262:14241-14249 Michell RH, Drummond AH, Downes CP: Inositol Lipids in Cell Signalling. London, Academic Press, 1989

J

systems

second-messenger

Dl

J 1990; 4:3178-3188 12.

messenger

229

1 . McGeer PL, Eccles JC, McGeer EG: Molecular Neurobiology of the Mammalian Brain, 2nd ed. New York, Plenum, 1987 2. North RA: Neurotransmitters and their receptors: from the clone to the clinic. Seminars in Neuroscience 1989; 1:81-90 3. Birnbaumer L: G proteins in signal transduction. Annu Rev Pharmacol Toxicol 1 990; 30:675-705 4. Gilman AG: G proteins: transducers of receptor-generated signals. Annu Rev Biochem 1987; 56:615-649 5. Nicoll RA, Malenka RC, Kauer JA: Functional comparison of neurotransmitter receptor subtypes in mammalian central nervous system. Nature 1989; 87:741-746 6. Ewald DA, Pang IH, Sternweis PC, Miller RJ: Differential G protein-mediated coupling of neurotransmitter receptors to Ca2+ channels in rat dorsal root ganglion neurons in vitro. Neuron 1989; 2:1185-1193 7. Bokoch GM, Katada T, Northup JK, Hewlett EL, Gilman AG: Identification of the predominant substrate for ADP-ribosylation by islet activating protein.J BiolChem 1983; 258:2072-2075 8. Sternweis PC, RobishawJD: Isolation oftwo proteins with high affinity for guanine nucleotides from membranes of bovine brain. J Biol Chem 1984; 259:13806-13813 9. Taylor SJ, Smith JA, Exton JH: Purification from bovine liver membranes of a guanine nucleotide-dependent activator of phosphoinositide-specific phospholipase C: immunologic identification as a novel G-protein alpha subunit. J Biol Chem 1990; 265:17150-17156 I 0. Freissmuth M, Casey PJ, Gilman AG: G proteins control diverse

FASEBJ

Second

HK, Simon MI, Gilman AG: Gz, protein with unique biochemical

J Biol Chem

Worley

27.

signaling.

SH:

Miller RJ: Multiple calcium channels Science 1987; 235:46-52 Neer EJ: Structural and functional studies Gen Physiol 5cr 1990; 45:143-15 1

on

REFERENCES

oftransmembrane

PF, Snyder

1990; 265:2383-2390 Linden J, Delahunty TM: Receptors tide breakdown. Trends Pharmacol

26.

pathways

Worley

Biol Chem

serves

ity

17.

disorders)

JM,

K. MANJI

37.

38.

39.

40.

41.

and

J, PremontJ, D2

of Clinical

Psychoneuroendocrinol-

Guilford Press, 1987 P, Drouva S, Rasolonjanahary

C, of adenylate cyclases by 17 betaneurons and anterior pituitary cells.

Enjalbert

dopamine-sensitive

estradiol in cultured striatal J Neurochem 1989; 52:410-418 Goldman-Rakic PS, Lidow MS. Galiager

paminergic, adrenergic, and serotoninergic plementarity of their subtypes in primate Neurosci 1990; 10:2125-2138 McCormick DA, Williamson A: Convergence neurotransmitter action in human cerebral Acad Sci USA 1989; 86:8098-8 102 Ross EM: Signal sorting and amplification coupled receptors. Neuron 1989; 3:141-152

Aghajanian GK, Wang YY: Common tor mechanisms in the locus coeruleus: brain North

slices. RA,

Neuropharmacology Williams JT:

On

R, Kordon

A: Differential

alpha

modulation

DW:

Overlap

receptors prefrontal

of doand comcortex. J

and divergence cortex. Proc through

of NatI

G protein-

2- and opiate

intracellular studies 1 987; 26:793-799 the potassium conductance

effecin in-

757

G PROTEINS

creased (Lond) 42.

43.

by

opioids

1985;

locus

coeruleus

J Physiol

neurones.

Andrade R, Malenka RC, Nicoll RA: A G protein couples serotonin and GABAB receptors to the same channels in hippocampus. Science 1986; 234:1261-1265 Lacey MG, Mercuri NB, North RA: On the potassium conductance

increase

activated

in rat substantia 437-453 44.

in rat

cology: The Third Generation HY. New York, Raven Press,

364:265-280

by GABAB

nigra

J

neurones.

and dopamine D2 receptors Physiol (Lond) 1988; 401:

pie to G proteins

B, SieverlingJ, on noradrenergic

and interact

with

Hertting nerve

64.

Nagy LE, Diamond I, Collier K, Lopez L, Ullman B, Gordon AS: Adenosine is required for ethanol-induced heterologous desensitization. Mochly-Rosen

the alpha

G: Presynaptic terminals cou-

2-adrenoceptors.

tion

66.

45.

46.

47. 48.

49.

50.

Si.

52.

53.

54.

Duman RS, Enna SJ: Modulation of receptor-mediated AMP production in brain. Neuropharmacology 1987;

26:98

986 Bouvier

Ann

57.

58.

59.

between

second

messengers.

C phosphorylates

the inhibitory

RM,

gene

ES, Spiegel

AM:

in Albright hereditary gradient gel electrophoresis.

Mutations

of the Gs alpha-

osteodystrophy Proc Natl

68.

.

69.

61.

Horn

Bilezkian JP: Mechanism of abnormal transmembrane signaling of the beta-adrenergic receptor in congestive heart failure. Circulation 1990; 80:271-283 Weinstein LS, Shenker A, Gejman PV, Merino MJ, Friedman E, Spiegel AM: Activating mutations of the stimulatory G protein in McCune-Albright syndrome. N EnglJ Med 1991; 325:1688-

72.

758

Shuckit

MA:

Biology

of risk

for alcoholism,

in Psychopharma-

Biochem

B, Estrin

W, Gordon

levels of cAMP

alcoholic

patients.

Nature

differentially

Biophys

Res

Corn-

Proc

A: Basal

and

are reduced

Natl

Acad

adenosine

in lymphocytes

Sci

USA

1987;

84:

Tabakoff B, Hoffman PL, Lee JM, Saito T, Willard B, Dc-Leon Jones F: Differences in platelet enzyme activity between alcoholics and nonalcoholics. N EnglJ Med 1988; 318:134-139 Nestler EJ, Erdos JJ, Terwilliger R, Durnan RS, Tallman iF: Regulation of G proteins by chronic morphine in the rat locus coeruleus. Brain Res 1989; 476:230-239

.

K, Beitner-Johnson EJ: Opiate withdrawal

DB, KrystalJH, Aghajanian and the rat locus coeruleus:

havioral, electrophysiological, and biochemical Neurosci 1990; 10:2308-2317 Attali B, Vogel Z: Long-term opiate exposure leads of the alpha i-i subunit of GTP-binding proteins.

GK, be-

J

correlates. to reduction

J Neurochem

1989; 53:1636-1639 Nestler EJ, Terwilliger RZ, Walker JR. Sevarino KA, Duman RS: Chronic cocaine treatment decreases levels of the G protein subunits

Gi alpha

J Neurochem

and

1990; M, Kleinman

Go

alpha

in discrete

55:1079-1082 JE, Hanbauer

regions

brain.

Memo

74.

recognition sites with adenylate cyclase in nuclei accumbens and caudatus of schizophrenics. Science 1983; 221:1304-1307 Seeman P, Niznik HB, Guan HC, Booth G, Ulpian C: Link be-

tween nia 75.

76.

77.

Dl and D2 dopamine and

Huntington

receptors

diseased

brain.

of dopamine

is reduced Proc

Kerwin RW, Beats BC: parahippocampal gyrus phrenic brain determined

Acad

Davies

(Gi,

Go)

brain.

AA: Mitral

hypersensitivity: with desensitization exposure. Am J Med 1987; 82:193-20 1 AO, Su CJ, Balasubramanyam A, Codina

L: Abnormal dysautonomia:

Sci

USA in the

J Neural

Increased forskolin binding in the left and CAl region in post mortem schizoby quantitative autoradiography.

Neurosci Lett 1990; 118:164-168 Davies AO, Mares A, PoolJL, Taylor

terenol

Dl

in schizophre-

Nati

1989; 86:10156-10160 Okada F, Crow TJ, Roberts GW: G-proteins basal ganglia of control and schizophrenic Transrn Gen Sect 1990; 79:227-234

with symptoms renergic receptor 78.

I: Coupling

of rat

73.

of beta-adrenergic supercoupling

guanine nucleotide regulatory evidence from reconstitution

valve prolapse beta 2-adon isopro-

J, Birnbaumer protein in MVP of Gs. J Clin En-

docrinol

79.

80.

81.

82.

1695 62.

I, Wrubel

Nestler

detected by Acad Sci USA

EM,

RNA.

M: Ethanol

cells.

I,

desensitiza-

s messenger

LA, Henteleff

Rasmussen

70.

1990; 87:8287-8290 60.

Diamond

1413-14 16

1986; 47:890-897

Gershon

subunit denaturing

alpha

in neural

M,

heterologous

1988; 155:138-143

from

guanine-nucleotide-

Sagi-Eisenberg R: GTP-binding proteins as possible targets for protein kinase C action. Trends Biochem Sci 1989; 14:355-357 Nishizuka T: The molecular heterogeneity of protein kinase C and its implications for cellular regulation. Nature I 988; 85: 661-665 Patten JL, Levine MA: Immunochemical analysis of the alphasubunit of the stimulatory G-protein of adenylyl cyclase in patients with Albright’s hereditary osteodystrophy. J Clin Endocrinol Metab 1990; 71:1208-1214 Weinstein LS, Gejman PV, Friedman E, Kadowaki T, Collins

causes

by reducing

receptor-stimulated

NY

binding regulatory component and apparently suppresses its function in hormonal inhibition of adenylate cyclase. Eur J Biochem 1985; 151:431-437 Olianas MC, Onali P: Phorbol esters increase GTP-dependent adenylate cyclase activity in rat brain striatal membranes. J Neu-

ethanol

G proteins

Diamond

1-

A2. Psychopharmacol Bull 1991; 27:247-253 Jakobs KH, Bauer S, Watanabe Y: Modulation of adenylate cyclase of human platelets by phorbol ester: impairment of the hormone-sensitive inhibitory pathway. Eur J Biochem 1985; 151:425-430 Katada T, Gilman AG, Watanabe Y, Bauer S, Jakobs KH: Pro-

rochem 56.

67.

cyclic

Acad Sci 1990; 594:120-129 Asakura M, Tsukamoto T, Kubota H, Osada K, Imafuku J, Nishizaki J, Sato A, Nakanishi J, Shimbo K, Shibata M: Involvement of protein kinase in the regulation of beta-adrenergic receptors by antidepressants. Int J Clin Pharmacol Res 1989; 9: 123-130 Manji HK, Chen G, Bitran JA, Potter WZ: Downregulation of beta adrenergic receptors in vitro involves PKC/phospholipase

tein kinase

55.

Cross-talk

of receptors

mun

71 M:

AS: Chronic

regulates

and guanine nucleotide bindFASEB J 1990; 4:2612-2622

Mol Pharmacol 1989; 36:744-748 D, Chang FH, Cheever L, Kim

1988; 333:848-850 Charness ME, Querimit

J

Neurochem 1989; 53:1629-1635 Adamson P, Mantzouridis T, Xiang JZ, Hajimohammadreza I, Brammer MJ, Campbell IC: Alpha-2-adrenergic, kappa-opiate, and P1-purinergic autoreceptors have mutually antagonistic effects: a new regulatory mechanism? J Neurochem 1989; 53: 1077-1082 Limberger N, Spath L, Starke K: Presynaptic alpha 2-adrenoceptor, opioid kappa-receptor and adenosine Al-receptor interactions on noradrenaline release in rabbit brain cortex. Naunyn Schmiedebergs Arch Pharmacol 1988; 338:53-61 Taylor CW: The role of G proteins in transmembrane signalling. BiochemJ 1990; 272:1-13 Rodbell M: Programmable messengers: a new theory of hormone action. Trends Biochem Sci 1985; 10:461-464

by Meltzer

Hoffman PL, TabakoffB: ing proteins: a selective

65.

Ethanol interaction.

Edited

63.

Gordon

Allgaier C, Daschmann kappa-opioid receptors

of Progress. 1987

83.

Metab 1991; 72:867-875 J, Dc Backer JP, Ebinger G, Vauquelin G: Coupling of Dl dopamine receptors to the guanine nucleotide binding protein Gs is deficient in Huntington’s disease. Brain Res I 989; 496:327-330 Dewar D, Horsburgh K, Graham DI, Brooks DN, McCulloch J: Selective alterations of high affinity E3Hlforskolin binding sites in Alzheimer’s disease: a quantitative autoradiographic

Dc Keyser

study. Brain Res 1990; 511:241-248 Nomura Y, Kitamura Y, Kawai M, Segawa T: Alpha 2-adrenoceptor-GTP binding regulatory protein-adenylate cyclase systern in cerebral cortical membranes of adult and senescent rats. Brain Young

Res 1986; 379:118-124 LT, Li PP, Kish Si, Siu KP, Warsh JJ: Postmortem cerebral cortex G, a-subunit levels are elevated in bipolar affective disorder. Brain Res 1991; 553:323-326 Schreiber G, Avissar S, Danon A, Belmaker RH: Hyperfunc-

Am

J

Psychiatry

1 49:6,

June

1992

HUSSEINI

84.

85.

86.

tional G proteins in mononuclear leukocytes of patients with mania. Biol Psychiatry 1991; 29:273-280 Pandey GN, Pandey SC, Davis JM: Peripheral adrenergic receptors in affective illness and schizophrenia. Pharmacol Toxicol 1990; 3:13-36 Mann JJ, Brown RP, Halper JP, Sweeney JA, Kocsis JH, Stokes PE, Bilezikian JP: Reduced sensitivity of lymphocyte beta-adrenergic receptors in patients with endogenous depression and psychomotor agitation. N Engi J Med 1985; 3 13:715-720

Ebstein

RP, Lerer

B, Shapira

B, Shemesh

Z, Moscovich

Kindler S: Cyclic AMP second-messenger signal in depression. BrJ Psychiatry 1988; 152:665-669

87.

Charney

DS, Innis

Platelet

alpha-2-receptor

in panic 107 88.

89.

90.

91.

disorder.

93.

94.

95.

96.

97.

RS, Woods

binding

and

cyclase 1989;

Berridge

MJ, Downes of lithium:

CP, Hanley a unifying

MR: Neural hypothesis.

99.

Cell

1989;

59:411-

101.

cerebral

Jpn J Psychiatry

cortex.

Okada

F, Tokumitsu

substrates

T: Effects

adenylate

Neurol

Y, Ui M: Possible

coupling

1988;

cyciase

in cerebral

involvement

cortices

in rat

42:858-860 of pertussis

(Gi, Go) in desipramine-induced of rats.

refractoriness J Neurochem

1988; 51:194-199 Yamaoka K, Nanba

T, Nomura S: Direct influence of antideGTP binding protein of adenylate cyclase in cell membranes of the cerebral cortex of rats. J Neural Transm 1988; 71:165-175 Duman RS, Terwilliger RZ, Nestler EJ: Chronic antidepressant regulation ofGsa and cyclic AMP-dependent protein kinase (abstract). Pharmacologist 1989; 31:182 Menkes DB, Rasenick MM, Wheeler MA, Bitensky MW: Guaon

nosine triphosphate activation of brain hancement by long-term antidepressant 1 10.

of desipramine

cyclase

adenylate

cyclase:

treatment.

en-

Science

1983; 219:65-67 Newman ME, Lerer B: Post-receptor-mediated increases in adenylate cyclase activity after chronic antidepressant treat-

ment: relationship

to receptor

desensitization.

Eur J Pharmacol

1989; 162:345-352 1 1 1.

Ozawa binding

H, Rasenick MM: Coupling of the protein Gs to rat synaptic membrane

is enhanced subsequent Mol Pharmacol 1989; 112.

Duman

RS,

Strada

to chronic 36:803-808 Si,

Enna

SJ:

antidepressant Effect

of

GTPcyclase

treatment.

imipramine

Goldberg H, Clayman P, Skorecki K: Mechanism of Li inhibition of vasopressin-sensitive adenylate cyclase in cultured renal epithelial cells. Am J Physiol 1988; 255(5, part 2):995-1 002 Manji HK, Bitran JA, Masana MI, Chen G, Hsiao JK, Risby

MV, Potter

WZ:

Signal

transduction

to the tricyclic

antidepressant

Manji Chronic

HK, Chen exposure

104.

Okada

F, Tokumitsu

1987;

gic

receptor-coupled

in vivo treatment 47:454-459

J

Y, Ui M: Desensitization

Psychiatry

adenylate

of rats with

cyclase

desipramine.

cortex

J Neurochem

June

1992

Gleiter

CH,

DeckertJ,

and repeated

tors, adenylate 1 16.

chem Huang

1989; RR,

Nutt

DJ, Marangos

cyclase, and the adenosine

Strader

CD: Identification

guanine

nucleotide-binding

118.

1 19.

120.

121.

122.

123.

after

1986;

PJ: Electroconvulsive

neuromodulatory ECS on the adenosine

52:641-646 Dehaven RN,

col 1990; 37:304-3 10 1 1 7. Anthony BL, Dennison

124.

1 49:6,

teins in rat cortex. EurJ Pharmacol 1990; 189:99-103 Nishida A, Kaiya H, Tohmatsu T, Wakabayashi S, Nozawa Y: Electroconvulsive treatment: effects on phospholipase C activity and GTP binding activity in rat brain. J Neural Transm Gen Sect

of single

of beta-adrener-

in cerebral

norepi-

and electroconvulsive shock attenuate betaand muscarinic cholinoceptor coupling to G pro-

shock (ECS) and the adenosine

49:282-289

GA, Bitran JA, Gusovsky F, Potter WZ: of C6 glioma cells to desipramine desensitizes beta-adrenoceptors, but increases KLJKH ratio. Eur J Pharmacol 1991; 206:159-162

brain

1990; 81:121-1 30 1 15.

desipra-

mine reduces the number of beta-adrenoceptors. Biochem Pharmacol 1986; 35:1899-1902 Fishman PH, Finberg JP: Effect of the tricyclic antidepressant desipramine on beta-adrenergic receptors in cultured rat glioma

C6 cells. J Neurochem

1 14.

modulation

by lithium: cell culture, cerebral microdialysis, and human studies. Psychopharmacol Bull 1991; 27:199-208 Sulser F: Antidepressant treatments and regulation of norepinephrine-receptor-coupled adenylate cyclase systems in brain. Adv Biochem Psychopharmacol 1984; 39:249-261 Honegger UE, Disler B, Wiesmann UN: Chronic exposure of

cells in culture

Carbamazepine adrenoceptor

on the rat

and

nephrine-coupled cyclic nucleotide generating system: alteration in alpha and beta components. J Pharmacol Exp Ther 1985; 234:409-414 Avissar S, Schreiber G, Aulakh CS, Wozniak KM, Murphy DL:

1 13.

administration

stimulatory adenylate

Chuang DM: Neurotransmitter receptors and phophoinositide turnover. Annu Rev Pharmacol Toxicol 1989; 29:71-110 Mork A, Geisler A: Effects of GTP on hormone-stimulated adenylate cyclase activity in cerebral cortex, striatum, and hippocampus from rats treated chronically with lithium. Biol Psychiatry 1989; 26:279-288 Mork A, Geisler A: The effects of lithium in vitro and cx vivo on adenylate cyclase in brain are exerted by distinct mechanisms. Neuropharmacology 1989; 28:307-311 Manji HK, Hsiao JK, Risby ED, Oliver J, Rudorfer MV, Potter WZ: The mechanisms of action of lithium, I: effects on serotoninergic and noradrenergic systems in normal subjects. Arch Gen Psychiatry 1991; 48:505-512 Risby ED, Hsiao JK, Manji HK, Bitran J, Moses F, Zhou DF, Potter WZ: The mechanisms of action of lithium, II: effects on adenylate cyclase activity and beta-adrenergic receptor binding in normal subjects. Arch Gen Psychiatry 1991; 48:513-524 Avissar S, Schreiber G, Danon A, Belmaker RH: Lithium inhibits adrenergic and cholinergic increases in GTP binding in rat

103.

Am

Y, Saito

adrenocorticotropin

human

102.

Hatta

receptor

419

ED, Rudorfer

100.

109.

H,

on

pressants

and developmen-

cortex. Nature 1988; 331:440-442 98.

107.

GR:

activity 98:102-

F, Ikeda

administration

of adenylate

108.

(Berlin)

Tsuchiya

toxin

DG,

SW, Heninger

adenylate

Psychopharmacology

106.

amplification

Lerer B, Bleich A, Bennett ER, Ebstein RP, Balkin J: Platelet adenylate cyclase and phospholipase C activity in posttraumatic stress disorder. Biol Psychiatry 1990; 27:735-740 Berridge MJ, Downes CP, Hanley MR: Lithium amplifies agonist-dependent phosphatidylinositol responses in brain and salivary glands. Biochem J 1982; 206:587-595 Hallcher LM, Sherman WR: The effects oflithium ion and other agents on the activity of myoinositol1 -phophatase from bovine brain. J Biol Chem 1980; 255:10896-10901 tal actions

92.

RB, Duman

105.

K. MANJI

Cheung

of allosteric protein

RL, Aronstam

AH,

system: effect Al and A2 recepuptake site. J Neuro-

Diehl

RE,

antagonists interactions.

Dixon

RA,

of receptorMol

RS: Influence

Pharma-

of volatile

anesthetics on muscarinic regulation of adenylate cyclase activity. Biochem Pharmacol I 990; 40:376-379 Parsons WJ, Ramkumar V, Stiles GL: Isobutylmethylxanthine stimulates adenylate cyclase by blocking the inhibitory regulatory protein, Gi. Mol Pharmacol 1988; 34:37-41 Mousli M, Bueb JL, Bronner C, Rouot B, Landry Y: G protein activation: a receptor-independent mode of action for cationic amphiphilic neuropeptides and venom peptides. Trends PharmacolSci 1990; 11:358-362 Moor M, Honegger UE, Weismann UN: Organospecific, qualitative changes in the phospholipid composition of rats after chronic administration of the antidepressant drug, desipramine. Biochem Pharmacol 1987; 37:2035-2039 Bruns RF, Fergus JH: Allosteric enhancement of adenosine Al receptor binding and function by 2-amino-3-benzoylthiophenes. Mol Pharmacol 1990; 38:939-949 Milligan G: Techniques used in the identification and analysis of function of pertussis toxin-sensitive guanine nucleotide binding proteins. Biochem J 1988; 255:1-13 Simonds WF, Goldsmith PK, CodinaJ, Unson CG, Spiegel AM: Gi2 mediates alpha 2-adrenergic inhibition of adenylyl cyclase in platelet membranes: in situ identification with G alpha C-terminal antibodies. Proc Natl Acad Sci USA 1989; 86:7809-78 13 Akita Y, Saito T, Yajima Y, Sakuma S: The stimulatory and

759

G PROTEINS

inhibitory cyclase roidism Spiegel

125.

guanine

nucleotide-binding

in erythrocytes type I. J Clin AM, Levine

ciency

of hormone

from

receptor-adenylate

cyclase resistance. GM:

127.

128.

stimulatory guanosine with pressure-overload 1988; 81:420-424 Koski G, Simonds WF,

of agonists

J Biol

Chem

coupling Prog

Biol

Res

analysis mutants.

Klee

WA:

Guanine

nucleotides

opiate

use

Studying

Several distinct and used to study G proteins

in Clinical Populations

G Proteins

complementary approaches in clinical populations. The

can range

be of

techniques developed in recent years to study G protein interactions with both receptors and effectors is described in detail in an excellent

tial

problem

cold

NAD

the method

GTP-dependent

of

(127),

though

not,

when

tional changes.

used

alone,

provide

changes occurring For this reason,

valuable

adjunctive aspects

structural

namide-adenine

information

in the pertussis

tools

for

about

investigating

of various

G proteins.

dinucleotide

(NAD)

functional By using

and

form.

in ADP-ribosylating In contrast,

G protein

G in the activated,

the inactive,

is ADP-ribosylated

mal susceptibility ribosylation has

760

only

undissociated

by pertussis

toxin.

example,

a comparison

used

ADPcon-

does

not

exist.

Al-

can be generated pertussis G1 (122).

adenylyl

G protein),

cholera

as probes

by

toxin

cyclase

and

forskolin

of For

activity

and G protein (which

in re-

analogues

acts

at G, and

G protein,

and

toxin

pertussis

of the signal

and

effector

transduction

pathway

may

be

because

of

G and to inactivate/un-

activate GTP

should,

diof

to be identoxin

binding,

agonist-induced

agonist-induced increase in GTPase in theory, serve as good indexes for G protein in the receptor-effector coupling being studied.

and

activity involveHow-

ever, a number of practical problems exist. First, the inability of GTP or its analogues to enter intact cells across the plasma membrane necessitates the use of membrane preparations. Second,

in a number

of systems,

the

basal

GTP

binding

and

GTPase activity are high, perhaps because signal-transducing G proteins represent only one group of GTP binding and

hy-

drolyzing

the

high

proteins.

nonspecific

increases.

abnor-

memand

allow for the dissection protein/effector system.

fluoride

in receptor,

Similarly,

to preclude

of the G proteins to toxin-catalyzed been shown to result from an abnormal

of the

agonists,

at the

abnormalities

dissociated of the

a G

the catalytic unit of adenylyl cyclase), and Mn2 (which rectly stimulates adenylyl cyclase) may allow identification

ment

the

of

a model

Certain pharmacologic agents the multicomponent receptor/G

form

Thus,

of

G protein but have allows for the de-

!ymphoma

such

GDP release,

[32Plnicoti-

lack a specific and effector(s)

S49

in-

transfer of its [32P]ADP-ribose moiety to membrane components, it is possible to obtain information about functional aspects of the G protein. In such investigations, [32P]ADP-ribosylation of membrane components can be used because both cholera and pertussis toxin preferentially catalyze the ADP-ribosylation of a given conformational state (dissociated/undissociated) of the G proteins. Cholera toxin is very

effective

cyc

their ability to constitutively couple G, respectively. Finally, agonist-induced

and

determining

excess

extraction of G, from a donor full reconstitution of -adrenergic

do

conforma-

absence of quantitative and cholera toxin are

a large

treating appropriate membranes with then examining the effects of exogenous

tified.

they

including

activity

cumbersome,

adenylyl cyclase inhibition (123). era recognize both the dissociated

However, since these antisa subunits and the corre-

that functional

of the

more

act

complex,

by

mixture.

adenylyl cyclase activity in this cell line. Such studies have been used to demonstrate G, abin pseudohypoparathyroidism (125, 126), heart and mitral valve prolapse (78). At present, a

(which

4y

overcome

detergent for the

to receptor

heterotrimeric

pertussis toxin ribosylcases, it may be possible

Cyc

sponse

undissociated,

in various

problems as a means of tissues. First,

possess cyclase

Thus, allows

the

choice for quantitating G protein subunits. Similarly, these peptide antibodies can be used as specific probes of G protein receptor-effector coupling. Thus, although human platelets contain G, G2, G13, G , and G, only G.2 appears to functionally couple to the a2-a#{228}renergic receptor and thereby mediate

sponding

potential

protein.

(124). brane

of

failure

yields the greatest conformational (and G proteins. Several

is clearly

be

reaction

receptor(s)

normalities

of researchers have generated specific antisera against peptides corresponding to the predicted amino acid of various G protein subunits.

antisera

are

to intracel-

variants of the S49 lymphoma cell line lack G, but -adrenergic receptors and catalytic units of adenylyl

equivalent”

specific

there

G proteins

membranes

termination

era and toxin-catalyzed ADP-ribosylation information about the quantity and therefore functional) state of specific

use of these

can

in the

using

reconstitution

The

in relay-

receptors

ADP-ribosylation

quantifying

appropriate

review by Milligan (122); only a brief overview of clinically applicable methods is presented here. The simultaneous use of immunoblotting techniques with specific antis-

groups synthetic sequence

abnormalities

Another functional probe is the so-called reconstitution assay, in which the G protein from the tissue in question is cxtracted and used to restore deficient adenylyl cyclase activity.

inhibit

receptors.

256:1536-1538

for

However,

toxin-catalyzed

and

with

extracellular

both cholera toxin and, in particular, ate more than one G protein. In some

Thus, 1. Methods

to correlate from

(1 13).

of

detecting

the APPENDIX

effectors

in the

and

signals

to separate the G protein a subunits electrophoretically. Second, [32PJNAD+ can be degraded by endogenous NAD-glycohydrolases, thereby altering substrate availability. This poten-

Ci: Decreased

to soluble

state

molecular

lular defi-

protein

Clin

Somatic genetic of unresponsive

SF, Homcy

ing

triphosphate binding protein in dogs left ventricular failure. J Clin Invest

and antagonists

1981;

formational

with

J Cell Physiol 1975; 85:611-620 Longabaugh JP, Vatner DE, Vatner

binding

of adenylate

pseudohypoparathyEndocrinol Metab 1985; 61:1012-1017 MA: Pseudohypoparathyroidism:

as a cause of hereditary hormone 1982; 97:327-340 Coffino P, Bourne HR, Tomkins ofcyclic AMP action: characterization

126.

proteins

patients

in

in the

GTPase

an This

the study

Thus,

accurate assay

activities

(122),

perhaps

membrane

J

to

be

more

toxin

greater

abundance

of their

“noise”

of receptor-mediated

to pertussis

and their greater

Am

sufficient

however,

linked

because

of instances,

generate

determination appears,

of receptors

plasma

majority

enzymatic

Psychiatry

effective

G proteins

capacity

1 49:6,

June

in the (128).

1992