204 ( 1991) 1#5- 107 8 1991 Elsevier Science Peblishers B.V. All rights resewed ~14-2999/9~/$03.5~ ADONIS 00142999910073hJ
European Jottrmzi of ~~ur~ta~o~o~,
EJP 20944
Shoti communication
Claire Mermet and Luc Quintin Received 23 May 1991, revised MS received 27 August 1991, accepted 3 September 1991
The dose-dependent reduction in catechol metabolism induced by an imidazoline witk a,-adrenoceptor agonist specificity, . . clomdmc, was assessed (EDS,,= 7 Fg.kg-’ i.v.1with in vivo voltammetty in the rostra1 ventrolateral medulla of rats kept under halothane anesthesia and strictly controlled circulatory and ventilatory observed in intact as well as in barodeafferented rats. Rostra1 ventrolateral
medulla; Voltammetry;
conditions. This reduction
Imidazoline; cu,-Adrenoceptor
1. Introduction Recently, cardiovascular neurons were identified in the rostra1 ventrolateral (RVL) medulla on the basis of their specific relationship to blood pressure and their spinal projections (review in Guyenet, 1990). With the help of clonidine, an imidazol~ne with ~~-adrenoceptor specificity, a subset of these cardiovascular neurons was classified as clonidine-sensitive (Sun and Guyenet, 1986). The low conduction velocities of these RVL neurons allows one to infer their adrenergic nature (Sun and Guyenet, 1986; Guyenet, 1990). A technique highly specific for the in vivo detection of catechols (Gonon et al., 1984) detects a signal which can be used as an index of catecholamine biosynthesis, or catechol metabolism, of adrenergic cell bodies in the RVL medulla (Gillon et al., 1990). This technique was used to study the effect of clonidine on the adrenergic component of central cardiovascular control.
2. materials and methods The methods used have been described elsewhere (Gonon et al., 1984; Quintin et al., 1989; Gillon et al., 1990; Milne et al., 1990). Male Sprague-Dawley rats (300-350 g) were housed under standard conditions (temperature: 21 + l”C, light: 7-19 h) with rat chow
Correspondence to: L. Quintin. Laboratory of Envirunmental physiology, Grange-Blanche School of Medicine, 69373 Lyon Cedex 03. France. Tel. 33.78,77.71.45,fax 33.78.77.73.58.
in catechol metabolism was
agonists; Clonidine; Adrenaline
and water freely available up to the experiment. Anesthesia was induced with halothane (induction: 5%, surgery: 1.25%, recording: 0.8%) and monitored with an end tidal halothane monitor (Siemens G~l20) throughout surgery and recording. Muscle relaxation was produced with metocuriue (400 pg.kg-‘, Lilly) after checking for appropriate analgesia. ~~ec~a~ic~i ventilation was established (f = 48 min- ‘, oxygen = 40%) through a tracheotomy to maintain an end-tidal CO, = 40 mm Hg. Rectal temperature was maintained at 37& 0.5”C. Saline was infused (5 rn~.kg-‘.b-‘~ through a femoral vein catheter. Mean arteria1 pressure (MAP) and heart rate (HR) were measured and recorded through a femoral catheter with a Bentley Trantec 800 transducer, a Gould pressure processor and a Gould 2600s recorder. Arterial blood gases were measured on an ABL30 Radiometer; the tidal volume was altered or sodium bicarbonate (100 ~11 was injected when necessary. In some experiments, bilateral deafferentation from the baroreceptors was performed by cutting the carotid sinus nerve, as well as the vagus and the superior laryngeal nerve where they exit from the nodose ganglion, leaving the sympathetic trunk intact. The effectiveness of deafferentation was assessed by observing the bradycardia following the injection of phenylephrine (10 pg.kg-’ i-v.). The rat was then set prone in the stereotaxic frame and surgery was performed. A treated (70 Hz, O-2.85 V, 20 S; DC, -0.8 V, 5 s; DC, + 1.5 V, 5 s) carbon fiber electrode (Gonon et al., 1984) was lowered vertically through the cerebellum at the following coordinates: incisor bar -3 mm below the interaural line, zero: lambda, P = -3-3.2 mm, I_ = 1.9 mm, depth = -9 mm. Differential normal
analysis of variance for repeated measures. A P < 0.05 indicated significance. Results for CA.OC are expressed as a percentage of the baseline value calculated by averaging the 5 absolute values of the catechol peak height vs. background current (Gonon et al., 1984) obtained during the last 10 min prior to the pharmacological challenge. For each experiment, the individual decrease was determined as the mean of the 4 scans during which the maximal decrease in CA.OC +vas noticed. The maximal decrease observed after a dose of 100 pg.kg-’ of i.v. clonidine was taken as the 100% effect. The maximal decrease for each experiment was normalized to this 100% effect. The normalized values were plotted using a sigmoid power model. The dose of clonidine necessary to obtain a 50% decrease in CA.OC (ED,,,) was determined graphically.
pulse voltammetry (Gonon et al., 19S4) was performed with the following parameters: scan rate: 5 mV.s-‘, scan tential: -240 mV to + 160 mV, pulse amplitude: 30-50 mV, pulse duration: 40 ms, prepulse duration: SO-120 ms. This technique allowed the recording every 2 min of a catechol oxidation current (CA.OC) as ak (1) appearing at +55 mV in a restricted area pm deep; the potentia! did not change from the beginning to the end of recording; (2) at the above mentioned coordinates, and (3) which disappeared rapidly following the administration of pargyline (75 mg.kg-” i.p.. n = 7) or cY-methyl-p-tyrosine (250 mg.kg - ’ i.p., n = 5) at the end of some experiments. The amplitude of the CA.OC was 0.96 f 0.09 nA (mean i S.E.M., n = 25). To identify the recording site, a lesion was made through the electrode (DC, +5 V, I5 s: and the brain was serially cut from the obex in 20 Frn slices and stained with cresyl violet (fig. 1A). After surgery. detection of the CA.OC and stabilization, baseline recordings were made for 30 min and were foXowed by bolus injection of saline (150 ;LI i-v.) or clonidine (Boehringer-Ingelheim) dissolved in a similar volume of vehicle. Each animal received only one test dose of either saline or clonidine. Results are expressed as means f S.E.M. and were analyzed with an
3. Results Saline (n = 4) did not modify CA.OC, MAP (baseline: 112 f 5 mm Hg) or HR (baseline: 385 f 9 b.min-‘1 (fig. lB, n.s. for all variables over the whole study period). Clonidine decreased the CA.OC in a dose-dependent manner, from 3 to 100 pg.kgei i.v. (maximal
__ -30
0
30 llmo @I$
CAOC (% bw~llne) 130 7
1
20
A0
60
80
100
._ -30
0
CLONIDINE lug/kg]
30
Time
(tnI$
Fig. 1. Dose-dependent effect of clonidine on the catechol oxidation current (CA.OC) recorded by in vivo voltammetry in the rostra1 ventrolateral medulla of the rat. (A) Cresyl violet staining of the recording site (black arrow) after destruction of the carbon fiber electrode at the end of the experiment. The center of the lesion was located 1400 pm rostra1 to the obex defined as the closing of the fourth ventricle. Calibration: 1 mm. The left part of the figure is adapted from the Paxinos and Watson atlas (interaural: - 2.8 mm). Abbreliarions. PCS: nucleus paragigantocellularis, IO: inferior olive, sp5: spinal trigeminal nerve, py: pyramidal tract, Amb: nucleus ambiguus. (B) Effect of i.v. saline (n = 4, white diamond) and clonidine 30 pg.kg-’ i.v. (n = 5, black diamond) on CA.OC (means+S.E.M.). (Cl Dose-response curve for clonidine on the CA.OC. Each empty square represents the mean maximal decrease observed for one individual experiment: each rat received only one dose of clonidine i.v. The experimental points for maximal inhibition were adjusted with a sigmoid power plot. The equation is y = -26.27+(103.1-26.27)/(1+ (4.97/x)1.304) (goodness of fit = IO-“). The EDs,, is 7 pg.kg-’ i.v. (D) effect of clonidine 10 pg.kg-’ IN. on CA.OC in intact (white diamond, n = 5) and deafferented (black diamond, n = 4) rats.
107
decrease to 89 zt 2, 65 k 5, 51 + 3, 47 t- 5% of baseline CA.OC for each dose, respectively, n = 5 rats for each dose). Clonidine IO pg.kg-’ i.v. given to intact (n = 5) or to deafferentated animals (n = 4) caused a similar reduction in CA.OC (fig. lD, n.s. between deafferented and intact rats).
4. Discussion These data demonstrate, in the rat, that there is a dose-dependent reduction in the meta~lism of catecholamines in an area next to the rostra1 pole of the inferior olive where the ~ate~holaminergic cell bodies are adrenergic (Gillon et al., 1990). Indeed, the current generated by the oxidation of catechols (CA.OC) represents an indirect index of the metabolism of catecholamines of adrenergic cell bodies (Gillon et al., 1990). Thus the present data demonstrate in vivo, in a biochemically specific and a dynamic manner, an inhibition by clonidine of the catechol metabolism within the central sympathetic generator itself. In this respect, the CA.OC recorded in the RVL medulla is modified by circulatory changes in a specific manner (Milne et al., 1990) and by antidromic stimulation from the intermediolateral cell column (Gillon et al., 1990). Thus, the reduction in CA.OC observed here may represent an inhibition pf adrenergic neurons involved in central cardiovascular control. In this respect, the present data agree with the demonstration of a selective reduction, following clonidine, of the electrical activity of slow conducting medullospinal sympathoexcitatory neurons (Sun and Guyenet, 19861, which are likely to be adrenergic (Guyenet, 1990). The small difference in the ED,,, observed here (7 pg.kg-‘1 and elsewhere (2.5 hg.kg-‘) (Sun and Guyenet, 1986) may be secondary to the fact that two different indices of cellular activity are considered: catechol metabolism vs. electrical activity of presumably adrenergic cell bodies. The inhibition of catechol metabolism and the finding that the effect of clonidine was independent of cardiovascular afferents are consistent with a similar result obtained in the caudal ventrolateral medulla with single-unit recordings (Kannan et al., 1986) and with the drop in blood pressure noticed after microir,‘ections of clonidine in the ventrolateral medulla (Bousquet et al., 1981). Taken together, these data agree with about the site of action for clonidine, namely a,-adrenergic autoreceptors Iocated on adrenergic cell bodies and/or dendrites in the rostra1 ventrolateral medulla. However, these findings leave two issues open: (1) the effect of clonidine may be linked primarily to its imidazole
structure and not to its affinity for a,-adrenoceptors (Bousquet et al., 1984); (2) part of the effect of clonidine may arise from an interaction with receptors located on non-catecholaminergic neurons (Lorez et al., 1983). Thus, the present model may be convenient to study, in vivo with high anatomical resolution, the mechanism of action of centrally acting antihypertensives drugs on the adrenergic component of the central cardiovascular control.
Supported by Boehringer-lngelheim France. Universite Claude Bernard-Biologie Humaine, Found&ion pour la Recherche M&ditale. Rigion RhBne-Alpes. Pr. Gharib is thanked for his interest and support. G. Deb@, R. Cespuglio, J. Frutoso, R. Hughson. F. Gonon, A. Maillet, L Paqueton. J.M. Pequignot and J.P. Viale are thanked for their help and critical advice.
References BOuSquet. P.. J. Feldman, R. Bloch and J. Schwartz, 1981, Ti;e nucleus reticularis lateralis: a region highly sensitive to cionidine. Eur. J. Pharmacol. 69, 389. Bousquet, P.. J. Feldman and J. Schwartz. 1984, Central cardiovascular effects of alpha adrenergic drugs: differences between catecholamines and imidazolines, J. Pharmacol. Exp. Ther. 230. 232. Gillon, J.Y.. F. Richard, L. Quintin, J.F. Pujol and B. Renaud, 1990. Pharmacological and functionnal evidence for extracellular 3.6 dihydroxyphenylacetic acid as an index of metabolic activity of adrenergic neurons: an in vivo voltammetric study in the rat rostra1 ventrolateral medulla, Neuroscience 37. 421. Gonon. F.. F. Navarre and M. Buda, 1984, In vivo monitoring of dopamine release in the rat brain with differential normal p&e voltammetry. Anal. Chem. 56. 573. Guyenet, P.. 1990, Role of the ventral medulla oblongata in blood pressure regulation, in: Central Regulation of Autonomic Functions, eds. A.D. Loewy and K.M. Spyer (Oxford University Press, New York) p. 145. Kannan. H., T. Osaka, M. Kasai, S. Okuya and H. Yamashita, 1986, Electrophysiological properties of neurons in the caudal ventolaterai medulla projecting to the paraventricular nucleus of the hypothalamus in rats. Brain Res. 376. 342. Lorez, H.P., D. Kiss, M. Da Prada and G. Hauesler, 1983, Effects of clonidine on the rate of noradrenaline turnover in discrete areas of the rat central neIvous system. Naunyn-Schmiedeb. Arch. Pharmacol. 323, 307. Milne. B., L. Quintin and J.Y. Gillon. 1990. Changes in catecholamine metabolism in the rostra1 ventrolateral medulla following halothane and nitroprusside-induced hypotension: an in vivo electrochemical study, Brain Res. 518. 143. Quintin, L., J.Y. Gillon, C.F., Saunier, G. Chouvet and M. Ghignone. 1989, Continuous volume infusion improves circulatory stability in anesthetized rats, J. Neurosci. Meth. 30. 77. Sun. M.K. and P. Guyenet, 1986, Effect of clonidine and gammaaminobutyric acid on the discharges of medulla-spinal SYmPathoexcitatory neurons in the rat, Brain Res. 368. I.
European Jottrmzi of ~~ur~ta~o~o~,
EJP 20944
Shoti communication
Claire Mermet and Luc Quintin Received 23 May 1991, revised MS received 27 August 1991, accepted 3 September 1991
The dose-dependent reduction in catechol metabolism induced by an imidazoline witk a,-adrenoceptor agonist specificity, . . clomdmc, was assessed (EDS,,= 7 Fg.kg-’ i.v.1with in vivo voltammetty in the rostra1 ventrolateral medulla of rats kept under halothane anesthesia and strictly controlled circulatory and ventilatory observed in intact as well as in barodeafferented rats. Rostra1 ventrolateral
medulla; Voltammetry;
conditions. This reduction
Imidazoline; cu,-Adrenoceptor
1. Introduction Recently, cardiovascular neurons were identified in the rostra1 ventrolateral (RVL) medulla on the basis of their specific relationship to blood pressure and their spinal projections (review in Guyenet, 1990). With the help of clonidine, an imidazol~ne with ~~-adrenoceptor specificity, a subset of these cardiovascular neurons was classified as clonidine-sensitive (Sun and Guyenet, 1986). The low conduction velocities of these RVL neurons allows one to infer their adrenergic nature (Sun and Guyenet, 1986; Guyenet, 1990). A technique highly specific for the in vivo detection of catechols (Gonon et al., 1984) detects a signal which can be used as an index of catecholamine biosynthesis, or catechol metabolism, of adrenergic cell bodies in the RVL medulla (Gillon et al., 1990). This technique was used to study the effect of clonidine on the adrenergic component of central cardiovascular control.
2. materials and methods The methods used have been described elsewhere (Gonon et al., 1984; Quintin et al., 1989; Gillon et al., 1990; Milne et al., 1990). Male Sprague-Dawley rats (300-350 g) were housed under standard conditions (temperature: 21 + l”C, light: 7-19 h) with rat chow
Correspondence to: L. Quintin. Laboratory of Envirunmental physiology, Grange-Blanche School of Medicine, 69373 Lyon Cedex 03. France. Tel. 33.78,77.71.45,fax 33.78.77.73.58.
in catechol metabolism was
agonists; Clonidine; Adrenaline
and water freely available up to the experiment. Anesthesia was induced with halothane (induction: 5%, surgery: 1.25%, recording: 0.8%) and monitored with an end tidal halothane monitor (Siemens G~l20) throughout surgery and recording. Muscle relaxation was produced with metocuriue (400 pg.kg-‘, Lilly) after checking for appropriate analgesia. ~~ec~a~ic~i ventilation was established (f = 48 min- ‘, oxygen = 40%) through a tracheotomy to maintain an end-tidal CO, = 40 mm Hg. Rectal temperature was maintained at 37& 0.5”C. Saline was infused (5 rn~.kg-‘.b-‘~ through a femoral vein catheter. Mean arteria1 pressure (MAP) and heart rate (HR) were measured and recorded through a femoral catheter with a Bentley Trantec 800 transducer, a Gould pressure processor and a Gould 2600s recorder. Arterial blood gases were measured on an ABL30 Radiometer; the tidal volume was altered or sodium bicarbonate (100 ~11 was injected when necessary. In some experiments, bilateral deafferentation from the baroreceptors was performed by cutting the carotid sinus nerve, as well as the vagus and the superior laryngeal nerve where they exit from the nodose ganglion, leaving the sympathetic trunk intact. The effectiveness of deafferentation was assessed by observing the bradycardia following the injection of phenylephrine (10 pg.kg-’ i-v.). The rat was then set prone in the stereotaxic frame and surgery was performed. A treated (70 Hz, O-2.85 V, 20 S; DC, -0.8 V, 5 s; DC, + 1.5 V, 5 s) carbon fiber electrode (Gonon et al., 1984) was lowered vertically through the cerebellum at the following coordinates: incisor bar -3 mm below the interaural line, zero: lambda, P = -3-3.2 mm, I_ = 1.9 mm, depth = -9 mm. Differential normal
analysis of variance for repeated measures. A P < 0.05 indicated significance. Results for CA.OC are expressed as a percentage of the baseline value calculated by averaging the 5 absolute values of the catechol peak height vs. background current (Gonon et al., 1984) obtained during the last 10 min prior to the pharmacological challenge. For each experiment, the individual decrease was determined as the mean of the 4 scans during which the maximal decrease in CA.OC +vas noticed. The maximal decrease observed after a dose of 100 pg.kg-’ of i.v. clonidine was taken as the 100% effect. The maximal decrease for each experiment was normalized to this 100% effect. The normalized values were plotted using a sigmoid power model. The dose of clonidine necessary to obtain a 50% decrease in CA.OC (ED,,,) was determined graphically.
pulse voltammetry (Gonon et al., 19S4) was performed with the following parameters: scan rate: 5 mV.s-‘, scan tential: -240 mV to + 160 mV, pulse amplitude: 30-50 mV, pulse duration: 40 ms, prepulse duration: SO-120 ms. This technique allowed the recording every 2 min of a catechol oxidation current (CA.OC) as ak (1) appearing at +55 mV in a restricted area pm deep; the potentia! did not change from the beginning to the end of recording; (2) at the above mentioned coordinates, and (3) which disappeared rapidly following the administration of pargyline (75 mg.kg-” i.p.. n = 7) or cY-methyl-p-tyrosine (250 mg.kg - ’ i.p., n = 5) at the end of some experiments. The amplitude of the CA.OC was 0.96 f 0.09 nA (mean i S.E.M., n = 25). To identify the recording site, a lesion was made through the electrode (DC, +5 V, I5 s: and the brain was serially cut from the obex in 20 Frn slices and stained with cresyl violet (fig. 1A). After surgery. detection of the CA.OC and stabilization, baseline recordings were made for 30 min and were foXowed by bolus injection of saline (150 ;LI i-v.) or clonidine (Boehringer-Ingelheim) dissolved in a similar volume of vehicle. Each animal received only one test dose of either saline or clonidine. Results are expressed as means f S.E.M. and were analyzed with an
3. Results Saline (n = 4) did not modify CA.OC, MAP (baseline: 112 f 5 mm Hg) or HR (baseline: 385 f 9 b.min-‘1 (fig. lB, n.s. for all variables over the whole study period). Clonidine decreased the CA.OC in a dose-dependent manner, from 3 to 100 pg.kgei i.v. (maximal
__ -30
0
30 llmo @I$
CAOC (% bw~llne) 130 7
1
20
A0
60
80
100
._ -30
0
CLONIDINE lug/kg]
30
Time
(tnI$
Fig. 1. Dose-dependent effect of clonidine on the catechol oxidation current (CA.OC) recorded by in vivo voltammetry in the rostra1 ventrolateral medulla of the rat. (A) Cresyl violet staining of the recording site (black arrow) after destruction of the carbon fiber electrode at the end of the experiment. The center of the lesion was located 1400 pm rostra1 to the obex defined as the closing of the fourth ventricle. Calibration: 1 mm. The left part of the figure is adapted from the Paxinos and Watson atlas (interaural: - 2.8 mm). Abbreliarions. PCS: nucleus paragigantocellularis, IO: inferior olive, sp5: spinal trigeminal nerve, py: pyramidal tract, Amb: nucleus ambiguus. (B) Effect of i.v. saline (n = 4, white diamond) and clonidine 30 pg.kg-’ i.v. (n = 5, black diamond) on CA.OC (means+S.E.M.). (Cl Dose-response curve for clonidine on the CA.OC. Each empty square represents the mean maximal decrease observed for one individual experiment: each rat received only one dose of clonidine i.v. The experimental points for maximal inhibition were adjusted with a sigmoid power plot. The equation is y = -26.27+(103.1-26.27)/(1+ (4.97/x)1.304) (goodness of fit = IO-“). The EDs,, is 7 pg.kg-’ i.v. (D) effect of clonidine 10 pg.kg-’ IN. on CA.OC in intact (white diamond, n = 5) and deafferented (black diamond, n = 4) rats.
107
decrease to 89 zt 2, 65 k 5, 51 + 3, 47 t- 5% of baseline CA.OC for each dose, respectively, n = 5 rats for each dose). Clonidine IO pg.kg-’ i.v. given to intact (n = 5) or to deafferentated animals (n = 4) caused a similar reduction in CA.OC (fig. lD, n.s. between deafferented and intact rats).
4. Discussion These data demonstrate, in the rat, that there is a dose-dependent reduction in the meta~lism of catecholamines in an area next to the rostra1 pole of the inferior olive where the ~ate~holaminergic cell bodies are adrenergic (Gillon et al., 1990). Indeed, the current generated by the oxidation of catechols (CA.OC) represents an indirect index of the metabolism of catecholamines of adrenergic cell bodies (Gillon et al., 1990). Thus the present data demonstrate in vivo, in a biochemically specific and a dynamic manner, an inhibition by clonidine of the catechol metabolism within the central sympathetic generator itself. In this respect, the CA.OC recorded in the RVL medulla is modified by circulatory changes in a specific manner (Milne et al., 1990) and by antidromic stimulation from the intermediolateral cell column (Gillon et al., 1990). Thus, the reduction in CA.OC observed here may represent an inhibition pf adrenergic neurons involved in central cardiovascular control. In this respect, the present data agree with the demonstration of a selective reduction, following clonidine, of the electrical activity of slow conducting medullospinal sympathoexcitatory neurons (Sun and Guyenet, 19861, which are likely to be adrenergic (Guyenet, 1990). The small difference in the ED,,, observed here (7 pg.kg-‘1 and elsewhere (2.5 hg.kg-‘) (Sun and Guyenet, 1986) may be secondary to the fact that two different indices of cellular activity are considered: catechol metabolism vs. electrical activity of presumably adrenergic cell bodies. The inhibition of catechol metabolism and the finding that the effect of clonidine was independent of cardiovascular afferents are consistent with a similar result obtained in the caudal ventrolateral medulla with single-unit recordings (Kannan et al., 1986) and with the drop in blood pressure noticed after microir,‘ections of clonidine in the ventrolateral medulla (Bousquet et al., 1981). Taken together, these data agree with about the site of action for clonidine, namely a,-adrenergic autoreceptors Iocated on adrenergic cell bodies and/or dendrites in the rostra1 ventrolateral medulla. However, these findings leave two issues open: (1) the effect of clonidine may be linked primarily to its imidazole
structure and not to its affinity for a,-adrenoceptors (Bousquet et al., 1984); (2) part of the effect of clonidine may arise from an interaction with receptors located on non-catecholaminergic neurons (Lorez et al., 1983). Thus, the present model may be convenient to study, in vivo with high anatomical resolution, the mechanism of action of centrally acting antihypertensives drugs on the adrenergic component of the central cardiovascular control.
Supported by Boehringer-lngelheim France. Universite Claude Bernard-Biologie Humaine, Found&ion pour la Recherche M&ditale. Rigion RhBne-Alpes. Pr. Gharib is thanked for his interest and support. G. Deb@, R. Cespuglio, J. Frutoso, R. Hughson. F. Gonon, A. Maillet, L Paqueton. J.M. Pequignot and J.P. Viale are thanked for their help and critical advice.
References BOuSquet. P.. J. Feldman, R. Bloch and J. Schwartz, 1981, Ti;e nucleus reticularis lateralis: a region highly sensitive to cionidine. Eur. J. Pharmacol. 69, 389. Bousquet, P.. J. Feldman and J. Schwartz. 1984, Central cardiovascular effects of alpha adrenergic drugs: differences between catecholamines and imidazolines, J. Pharmacol. Exp. Ther. 230. 232. Gillon, J.Y.. F. Richard, L. Quintin, J.F. Pujol and B. Renaud, 1990. Pharmacological and functionnal evidence for extracellular 3.6 dihydroxyphenylacetic acid as an index of metabolic activity of adrenergic neurons: an in vivo voltammetric study in the rat rostra1 ventrolateral medulla, Neuroscience 37. 421. Gonon. F.. F. Navarre and M. Buda, 1984, In vivo monitoring of dopamine release in the rat brain with differential normal p&e voltammetry. Anal. Chem. 56. 573. Guyenet, P.. 1990, Role of the ventral medulla oblongata in blood pressure regulation, in: Central Regulation of Autonomic Functions, eds. A.D. Loewy and K.M. Spyer (Oxford University Press, New York) p. 145. Kannan. H., T. Osaka, M. Kasai, S. Okuya and H. Yamashita, 1986, Electrophysiological properties of neurons in the caudal ventolaterai medulla projecting to the paraventricular nucleus of the hypothalamus in rats. Brain Res. 376. 342. Lorez, H.P., D. Kiss, M. Da Prada and G. Hauesler, 1983, Effects of clonidine on the rate of noradrenaline turnover in discrete areas of the rat central neIvous system. Naunyn-Schmiedeb. Arch. Pharmacol. 323, 307. Milne. B., L. Quintin and J.Y. Gillon. 1990. Changes in catecholamine metabolism in the rostra1 ventrolateral medulla following halothane and nitroprusside-induced hypotension: an in vivo electrochemical study, Brain Res. 518. 143. Quintin, L., J.Y. Gillon, C.F., Saunier, G. Chouvet and M. Ghignone. 1989, Continuous volume infusion improves circulatory stability in anesthetized rats, J. Neurosci. Meth. 30. 77. Sun. M.K. and P. Guyenet, 1986, Effect of clonidine and gammaaminobutyric acid on the discharges of medulla-spinal SYmPathoexcitatory neurons in the rat, Brain Res. 368. I.
