Brain Research, 593 ( 19921311-313

311

Elsevier SciencePublishers B.V. BRES 25377

Chronic fluoxetine treatment reduces hypothalamic vasopressin secretion in vitro M a r g a r e t A l t e m u s a, G i o v a n n i C i z z a b a n d Philip W . G o l d b "Laboratory of Clinical Science, Dicision of b:tramural Research Programs. National institute of Mental Health, Bed~esda. MD 20892 (USA) and b Clinical Neuroendocrinology Branch, Division of b~tramural Research Programs. National b~stitute of Mental llc~lth. Bethesda. MD 20892 (USA)

(Accepted 30 June 19921

Key. words: Va~pressin; Fluoxetine; Hypothalamas;Obsessive-compulsivedisorder

Patients with obsessive-compulsivedisorder (OCD) hypersecrete the arousal producing neurohormone arginine vasopressin (AVP) into the cerebrospinal fluid and plasma. Because OCD responds preferentially to potent serotonin uptake inhibitors, we compared the effectof chronic fluoxetine treatment to that of other antidepressants (Irazodone and desipramine) on AVP release from rat hypothalamic organ culture and showed that only fluoxelinesignificantlyreduced in vitro AVP release.

Cerebrospinal fluid (CSF) and plasma arginine vasopressin (AVP) levels are significantly elevated in patients with obsessive compulsive disorder (OCD) t. This finding is of interest in the light of data that AVP administration to experimental animals delays the extinction of behaviors acquired during aversive conditioning Is, a phenomena that can be analogized to the obsessive's preoccupation with the potentially adverse consequences of a wide range of thoughts or actions. Obsessive.compulsive disorder is often accompanied by depressive symptoms, but is also characterized by recurrent, intrusive thoughts that often involve the fear that some potential danger has been left unchecked or that one is about to perform an act that is harmful to the self or others. Compulsive, stereotyped, repetitive behaviors or cognitions such as handwashing, hoarding or counting are often conducted to magically forestall the imagined danger and to relieve anxiety. And although many antidepressants are effective for treatment of major depression, only those which are selective or semi-selective serotonin reuptake inhibitors are also effective treatments for obsessive-compulsive disorder. 12(review) TO investigate the relevance of this neuropeptide abnormality in OCD to the efficacy of particular an-

tidepressant drugs for treatment of OCD, we evaluated the effects of chronic treatment with a variety of antidepressants on AVP secretion by single rat hypothalamic explants. Pharmacologic agents used for chronic treatment included fluoxetine, a bicyclic serotonin reuptake blocker known to be effective tot treatment of OCDI2; trazodone, a triazolopyridine derivative and a selective but weaker serotonin reuptake blocker which is apparently not effective for treatment of OCD ~; and desipramine, a norepinephrine selective trieyclic, also shown to be ineffective for treatment of OCD ~2. Male Sprague-l,~awley rats (175-225 g, Taconic Farms Silver Spring, MD) were administered saline or 5 mg/kg of desipramine, trazodone, or fluoxetine i.p. daily for 8 weeks. Desipramine and trazodone were obtained from Sigma (St. Louis, MO), and fluoxetinc from Eli Lilly (Indianapolis IN). The experiment was conducted on two consecutive days. Hypothalami from half of the rats in each drug treatment group were tested on each of the 2 days. The last drug dose was administered at 08.00 h and rats were sacrificed by decapitation between 10.30 and 11.00 h. The order of sacrifice was randomized according to the type of drug treatment. Hypothalamic blocks were rapidly removed with fine-point-curved scissors using a

Correspondence: M. Altemus, Building 10, Room 3D41 9000 RockvillePike, Bethesda, MD 20892, USA.

312 sterile technique, from the optic chiasm anteriorly, to the mammillary bodies posteriorly, and along the lateral hypothalamic sulci laterally. The depth of the fragments was about 3 ram. Immediately after explanration, the hyopthalami (one hypothalamus per well, 48-multiwell plate, Costar, Cambridge, MA) were placed in a water-jacketed incubator at 37°C, under a 95% Oe 5% CO 2 atmosphere for a 2 h preincubation. Medium M 199 with modified Earle's salt (Gibco, Grand Island, NY) containing 0.1% BSA (fraction V, Sigma, St. Louis, M e ) and 20uM bacitracin (Aldrich, Milwaukee, WI) was used for both preincubation and secretion measures. After preincubation, each hypothalamic block was passed through four successive wells, at 20 minute intervals. The hypothalami were transferred from one well to another using a nylon mesh grid (3 × 3 mm, Small Parts, Miami, FL). In the first three wells, spontaneous secretion was assesed by exposing the explants to plain medium. In the last well, depolarization-stimulated secretion, as well as tissue viability, were evaluated by exposing the hypothalami to 60 mM KCI. AVP was measured directly without extraction by a previously described radioimmunoassay technique I but using a different antibody (Arnel, New York). Final dilution of the antibody was 1:90,000. Sensitivity (ED,s)) of the assay was 2 pg/mi. Total binding was 25%. The intraassay coefficient of variation was 1% at 23 pg/ml and 2.4% at 49 pg/ml, Results are expressed as mean ± S.E.M. One.way repeated measures ANOVA was applied to tile three baseline values to determine whether baseline values were affected by order of collection. Two.way repeated-measures ANOVA followed by a-priori contrasts was used to evaluate the effects of chronic drug treatment and KCI stimulation on hypothalamic AVP release. The three basal secretion values were not affected by order of collection: (F., 7.~= 0.74, P = 0.48). Hypothalamic AVP release was significantly increased by stimulation with 60 mM KCI after basal

TABLE I

Effect of chroaic drug treatmem on basal and stimuh)ted hypothalamic secretion of A VP (pg / 0,4 ml / 20 n)itl) Mean of values from 3 wells.

Saline (n = 7) Desipramine (n -= 4) Trazodone (n = 6) Fluoxetine (n = 8) *

A VP.basal

A VP-stimulated

110 + 2{) gT± 13 142 + 7 60± 15

149 ± 25 169±50 162 + 41 93±23

* Significantly reduced compared Io saline ( P < 0.02).

secrection collections had been completed. (97+ 8 pg/0.4 ml/20 min (mean of 3 basal collections) vs. 137 +_ 16 pg/0.4 ml/20 min). (F3,63 -- 3.58, P < 0.02). The effect of chronic drug treatment on mean basal secretion values from hypothalami incubated in plain medium as well as values obtained after incubation with 60 mM KCI are shown in Table I. Chronic drug treatment did produce significant differences in AVP secretion (F3.63 -- 4.97, P < 0.01). AVP values after fluoxetine treatment were significantly reduced compared to saline (P < 0.02), while AVP secretion after trazodone and desipramine treatment did not differ from saline (trazodone: P -- 0.23; desipramine: P = 0.50). There was no significant interaction of drug treatment with stimulated secretion of AVP (Fo,~3 -- 0.96, P -- 0.8), indicating that tissue viability was not affected by particular drug treatments and suggesting that the maximal releasable pool of peptide was not affected by any of the chronic drug treatments. Reducc~ basal AVP secretion after fluoxetine treatment does not seem to be due to reduced tissue viability since KCI administration caused equivalent stimulation of AVP release. This assessment of tissue viability has been validated previously for CRH secretion via dye exclusion studies and electron microscopic examination 4. The mechanism by which chronic administration of a serotonin reuptake blocker might lower AVP secretion remains unclear, Acutely, central administration of serotonin or serotonin agonists stimulates central and pituitary AVP release in a number of experimental paradigm~ IH4. In addition, acute serotonin depletion impairs hypothalamie AVP release in response to cold "~ and osmotic 7,1~ stress in vivo. Because the chronic administration of fluoxetine seems to increase the fir. ing rate of serotonergic neurons s,ls, we postulate that this drug may produce down-regulation of serotonin receptors or postsynaptie processes that mediate hypothalamic AVP secretion. However, we cannot rule out mechanisms that are independent of fluoxetine effects on serotonergic neurotransmission. The relevance of changes in hypothalamic peptide secretion to CSF peptide secretion remain to be determined. It is unclear whether hypothalamic AVP secretion is a significant determinant of CSF levels or whether release from the hypothalamus and CSF release are regulated in parallel. Most of the hypothalamic AVP measured in this study is likely to come from the supraoptic nucleus which is located relatively superficially and where AVP turnover seems to be most rapid ~. However, there is evidence from lesion studies in rats ~ that most of the AVP in the CSF comes from the superchiasmatic nucleus of the hypothalamus.

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We cannot definitely determine whether the capacity of fluoxetine to reduce hypothalamic AVP secretion is of relevance to its preferential efficacy in the treatment of obsessive-compulsive disorder. In this regard, neurohormones other than AVP are present in elevated concentrations in the CSF of patients with OCD (e.g. CRH ~ and somatostatin2), and our preliminary data indicate that fluoxetine does not influence their release from hypothalamic organ culture. However, the central administration AVP has been shown to influence a number of cognitive processes and behaviors that may be of relevance to the syndrome of obsessivecompulsive disorder. First, intracerebroventricular administration of AVP is arousal producing and enhances responses to external stimuli t5 in a manner that may be analogous to the intense anxiety characteristic of obsessive-compulsive disorder. Second, intracerebroventricular administration of AVP, in rats significantly prolongs the maintenance of aversively conditioned behaviors ~5 which again may be analogous to obsessions and compulsions in humans, because these can be understood as highly conserved conditioned responses to events associated with anxiety 6. Third, in humans, the intranasal administration of AVP seems to narrow the focus of attention s, an effect that may be analogous to the focused range of intense concerns and repetitive actions in the patient with obsessive-compulsive disorder. Finally, the central administration of AVP to e~:perimental animals promotes grooming behaviors t° which resemble a number of common compulsions including handwashing, cleaning and trichotilIomania (chronic hair pulling). I Altemus, M., Pigott, T., Kalogeras, K., Demitrack, M., Dubbert, B., Murphy, D.L. and Gold, P.W., Abnormalities in the regulation of vasopressin and corticotropin releasing factor secretion in obsessive-compulsive disorder, Arch. Gt,n, Psychiatry, 49 (1992) 9-20,

2 Altemus, M., Pigott, T., L'Heureux, F., Davis, C., Rubinow, D., Murphy, D.L. and Gold, P.W., CSF somatostatin in obsessivecompulsive disorder, Am. J. Psychiatry, in press.

3 Angelucci, L. and Scaccianoce, S., Mechanisms in the control of stress responsiveness, Ann. 1st Super Sanita., 26 (1990) 75-8. 4 Calogero, A.E., Bernardini, R., Margioris, A.N., Bagdy, G., Gailucci, W.T., Munson, P.J., Tamarkin, L., Tomai, T.P., Brady, L., Gold, P.W. and Chrousosm G.P., Effects of serotonergic agonists and antagonists on corticotropin-releasing hormone secretion by explanted rat hypothalami, Peptides, 10 (1989) 189-200. 5 deMontigny, C., Chaput, Y. and Blier, P., Modification of serotonergic neuron properties by long-term treatment with serotonin reuptake blockers, J. Clin. Psychiatry, 51 Suppl. (1990) B: 4-8. 6 Foa, E.B., Steketee, G.S. and Ozarow, BJ., Behavior therapy with obsessive-compulsives. In M. Mavissakalian (Ed.) OCD." Psychological and Pharmaacologicai Treatments, Plenum, New York, NY. 1985, pp. 49-70. 7 lovino, M. and Steardo, L., Effect of substances influencing brain serotonergic transmission on plasma vasopressin levels in the rat, Eur. J. Pharmacol., 13 (1985) 99-103. 8 Jennings, J.R., Nehes, R.D. and Reynolds, C.F., Vasopressin peptide may narrow the focus of attention in normal elderly, Psychiatry Res., 17 (1986) 31-39. 9 Liu, B., Kwok, R.P. and Fernstrom, J.P., Colchicine-induced increases in immunoreactive neus'opeptide levels in the hypothalamus: use as an index of bios~Jithesis, Life Sci., 49 (1991) 345-52. 10 Meisenberg, G. and Simmons, W.H., Behavioral effects of intracerebroventriculay administered neurohypophyseal hormone analogs in mice, Pharmacol. Biochem. Behat'., 16 (1982) 819-825. 11 Montes, R. and Johnson, A.K., Efferent mechanisms mediating renal sodium and water excretion induced by centrally administered serotonin, Am. J. Physiol., 259 (1990) RI267-73. 12 Murphy, D.L., Pato, M.T. and Pigott, T.A., Obsessive-compulsive disorder: treatment with serotonin-selective uptake inhibitors, asapirones and other agents, J. Clin. Psychopharmacol., 10 (1990) 91S-100S. 13 Pigott, T.A., L'Heureux, F., Rubenstein, C.S., Bernstein, S.E., Hill, J.L. and Murphy, D.L., A double-blind placebo controlled study of trazodone in patients with obsessive-compulsive disorder, J. Clin. Psychopharmacol., in press. 14 Pergola, P.E. and AIper, R.H., Vasopressin and autonomic mechanisms mediate cardiovascular actions of central serotonin, Am. J. Physiol., 260 (1991) R1188-RI 193. 15 Sahgal, A., A critique of the vasopressin-memory hypothesis, P,vychophurnaacolo~, 83 (1984) 215-228. I• Schwartz, WJ. and Reppert, S.M, Neural regulation of the circadian vasopressin rhythm in ccrebro spinal fluid: a prc.cminent role for the superchiasmatic nuclei, J. Neurosci., 5 (1¢)85) 2771-2778, 17 Stein, J.M., Lind, R.W. and Johnson, A.K., Central scrnloncrgic influences on renal electrolyte and water excretion, Ne, rophar. macology, 26 (1987) 1685-1692. 18 Wong, D.T., Reed, L.R., Bymaster, F.P. and Threlkeld, P.G., Chronic effects of fluoxetine, a selective inhibitor of serotonin uptake, on neurotransmitter receptors. J. Neural Tra,sm., 64 (1985) 251-269.

Chronic fluoxetine treatment reduces hypothalamic vasopressin secretion in vitro.

Patients with obsessive-compulsive disorder (OCD) hypersecrete the arousal producing neurohormone arginine vasopressin (AVP) into the cerebrospinal fl...
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