P-chlorophenylalanine Methyl Ester Research Articles

Determination of plasma mexiletine levels with gas chromatography—mass spectrometry and selected-ion monitoring

Mexiletine, 1-(2,6-dimethylphenoxy)-2-aminopropane (Mexitil), is an orally effective agent useful in the treatment of serious ventricular arrhythmias. This paper describes a gas chromatographic—mass spectrophotometric assay with selected-ion monitoring for the measurement of plasma or serum mexiletine levels. The drug and internal standard ( p-chlorophenylalanine methyl ester) were extracted from plasma into ethyl acetate—hexane—methanol (60:40:1, v/v). After separation and evaporation of the organic phase, the drug and internal standard were derivatized to their pentafluoropropyl derivatives prior to analysis. The reproducibility of the daily standard curve yielded mean inter- and intra-day coefficients of variability from 0.7 to 11.0%. The coefficients of variability for control plasma samples (0.5 and 1.0 μg/ml) ranged from 2.6 to 5.0% and the accuracy of the assay was 106 ± 6 and 100 ± 5% for the low and high level controls respectively. The limit of quantitation for the assay was 0.1 μg/ml. No interfering peaks were detected in any patient samples. This method can be used as a primary analytical method to measure mexiletine plasma levels or can serve as a convenient back-up method to HPLC procedures when contaminating peaks coelute with mexiletine.

Serotonin depletion by p-chlorophenylalanine decreases VIP mRNA in the suprachiasmatic nucleus.

The influence of serotonergic inputs on the levels of vasoactive intestinal peptide (VIP) mRNA in the rat suprachiasmatic nucleus (SCN) was examined by in situ hybridization combined with morphometrical analysis. Depletion of serotonin by p-chlorophenylalanine methyl ester (300 mg/kg i.p. daily for 3 days) caused a marked decrease in VIP mRNA signal levels in the ventrolateral part of the SCN. This finding suggests that serotonergic input controls VIP production at the mRNA level.

Involvement of dopamine D 2 receptor mechanism in the REM sleep deprivation-induced increase in swimming activity in the forced swimming test

Effects of monoamine synthesis inhibitors and dopamine antagonists on rapid eye movement sleep (REMs) deprivation treatment-induced increase in swimming activity were examined. Mice were deprived of REMs for 48 h by a small pedestal method. Swimming activity in REMs-deprived mice was significantly higher than those in group-housed or socially isolated animals used as the control. dl- α-Methyl- p-tyrosine methyl ester HCl (250 mg/kg, IP) decreased the swimming activity in REMs-deprived mice, whereas neither disulfiram (400 mg/kg, SC), a noradrenaline synthesis inhibitor, nor dl- p-chlorophenylalanine methyl ester HCl (300 mg/kg, IP) changed it. (+)-SCH23390 HCl (30 and 100 gmg/kg, IP), a selective dopamine D 1 antagonist, did not affect the activity in REMs-deprived mice. (±)-Sulpiride (12.5 and 25 mg/kg, IP), a selective dopamine D 2 antagonist, dose-dependently decreased swimming activity in REMs-deprived mice, while it did not significantly change the swimming activity in the control animals. These results suggest that REMs deprivation treatment-induced increase in swimming activity is mainly due to the functional changes in the dopaminergic system rather than the noradrenergic or serotonergic system, and that dopamine D 2 but not D 1 receptor mechanism is involved in the increase in swimming activity in REMs-deprived animals.

Mechanisms of l-N G-nitro arginine methyl ester-induced antinociception in mice: A role for serotonergic and adrenergic neurons

1. 1. The mechanisms involved in the antinociceptive action of l-N G-nitro arginine methyl ester ( l-NAME) were investigated in mice. 2. 2. Intraperitoneal administration of l-NAME produced a dose-dependent antinociception in the tail-flick, hot-plate and phenyl- p-quinone-induced writhing tests. 3. 3. Pretreatment with the catecholamine depletors 6-hydroxydopamine (5 μg i.c.v.) or reserpine (5 mg/kg i.p.) or the serotonin synthesis inhibitor, p-chlorophenylalanine methyl ester (200 mg/kg i.p. on 2 consecutive days) resulted in a significant decrease in the antinociceptive effect of l-NAME. 4. 4. Similarly, pretreatment with the selective α 1-adrenoceptor antagonist prazonin (2.5 mg/kg, i.p.), or the non-selective α-adrenoceptor blocker, phentolamine (5 mg/kg, i.p.) antagonized the antinociceptive effect of l-NAME. 5. 5. However, the administration of the selective α 2-adrenoceptor antagonist, idazoxan (2.5 mg/kg i.p.) was without effect. 6. 6. Likewise, pretreatment with the serotonin 5-HT2 receptor blocker, ketanserin (1 mg/kg, i.p.), the D2 dopamine receptor antagonist (±) sulpiride (30 mg/kg, i.p.) or the opioid antagonist naloxone (5 mg/kg,i.p.) did not inhibit the antinociceptive effect of l-NAME. 7. 7. These results suggest that l-NAME produces antinociception in the mouse probably by an action on adrenergic and serotonergic synapses.

Interaction between Norepinephrine and Serotonin in the Neuroendocrine Control of Growth Hormone Release in the Rat*

Both alpha 2-adrenergic (alpha 2) and serotonergic (5HT) neurons are associated with stimulation of GH secretion via GRH release. The object of this study was to determine whether the 5HT system is involved in the stimulation of GH secretion by alpha 2-receptor agonists. There are two parts of this study. In the first, the relationship between alpha 2-5HT systems were analyzed by determining if alpha 2-stimulated GH release is mediated by 5HT. In this model, systemically administered alpha 2-agonists [clonidine (CLON) or UK14,304] were tested against 5HT antagonists (meterogoline or cyproheptadine) or 5HT synthesis inhibitors (p-chlorophenylalanine methylester hydrochloride). In the second, sites of 5HT-GRH interaction were determined by testing the response to CLON after 5HT neurotoxin [5,7-dihydroxytryptamine (5,7-DHT)] microinjection at specific hypothalamic nuclei. In both experiments sequential blood samples were withdrawn from silastic jugular cannulas in unanesthetized, freely moving animals. Metergoline (0.045 and 0.135 mg/kg, iv) and cyproheptadine (0.969 micrograms/kg, iv) suppressed, in a dose-dependent manner, the CLON (33 or 66 micrograms/kg, iv)-induced GH surge that was detected 15-30 min after injection in control animals. Both cyproheptadine (0.969 micrograms/kg, iv) and p-chlorophenylalanine (300 mg/kg, ip) effectively suppressed the UK14,304 (220 micrograms/kg, iv)-induced GH surge that occurred 15-30 min after injection in control animals. These data suggest that an intact 5HT system is required for alpha 2-stimulated GH release. 5,7-DHT neurotoxin microinjected into the midline arcuate nucleus (6 micrograms/mg,iv) or bilaterally into the ventromedial nucleus or perifornical area (4 micrograms/0.2 microliter) 5 days previously suppressed the CLON (30 micrograms/kg)-induced GH surge only in animals with arcurate nucleus lesions. To determine if the suppression was mediated by inhibition of GH-releasing hormone (GRH) or stimulation of somatostatin (SRIF), an additional experiment was conducted including 5,7-DHT arcuate nucleus-lesioned animals injected with anti-SRIF. Inasmuch as anti-SRIF failed to reverse the 5,7-DHT suppression of GH secretion, the results of this experiment suggest that GRH mediates NE-5HT-induced GH secretion. In conclusion, these data suggest that alpha 2 activation of GH secretion requires intact serotonergic terminals in the arcuate nucleus and most likely involves GRH rather than SRIF, release.

Stress-induced prolactin release in female, male and androgenized rats: influence of progesterone treatment.

Ether stress applied at 10.00 h induced a 100% increase in serum prolactin in intact and ovariectomized androgenized rats. Ovariectomy significantly diminished the basal serum prolactin values observed in intact androgenized rats. Two doses of progesterone (5 mg) given to intact and ovariectomized androgenized rats 14 and 2 h before exposure to ether stress increased prolactin values in the control groups but completely prevented the effect of stress. Exposure to ether stress induced a 100% increase in serum prolactin values in androgenized rats with increased serum progesterone levels 4 days after the induction of ovulation and the luteal phase with human chorionic gonadotropin (hCG). A group of androgenized rats with induced maternal behaviour and which had been suckled for 6 days was given 100 i.u. hCG and suckled for another 6 days after the hCG-induced luteal phase had been established. The serum prolactin and progesterone values of these rats were significantly higher than those treated with hCG only and ether stress did not increase prolactin release. A greatly increased serum concentration of prolactin was obtained in pro-oestrous and oestrous virgin rats after exposure to ether stress. Serum prolactin was also increased by stress in male rats. Progesterone administration to these female and male rats prevented stress-induced prolactin release. To ascertain the part played by dopamine and serotonin in the effect of stress on prolactin release, groups of androgenized and oestrous female rats were treated with bromocriptine or p-chlorophenylalanine methylester hydrochloride (pCPA).(ABSTRACT TRUNCATED AT 250 WORDS)

Serotonin and norepinephrine-dependent effects of fenfluramine on plasma renin activity in conscious male rats

Administration of dl-fenfluramine to male rats caused an initial rise, followed by a sustained decrease in plasma renin activity. Both the increase, which reached a maximum at 30 min and the decrease, which was maximal at 4 hr after administration of fenfluramine, were dose-dependent. Pretreatment with either of the blockers of serotonin uptake, fluoxetine or indalpine blocked the increase in plasma renin activity induced by fenfluramine at 30 min, but did not affect the decrease at 4 hr after injection. Similarly, pretreatment with the inhibitor of the synthesis of serotonin, p-chlorophenylalanine methylester (PCPA) blocked the initial (30 min) but not the delayed (4 hr) effect of fenfluramine on plasma renin activity. The initial stimulation of secretion of renin by a submaximal dose (2 mg/kg, i.p.) of fenfluramine was potentiated by pretreatment with the precursor of serotonin l-tryptophan (100 mg/kg, i.p.). Pretreatment with the blocker of the uptake of norepinephrine, desipramine did not prevent the initial (30 min) effect but completely prevented the delayed (4 hr) effect of fenfluramine on plasma renin activity. These results suggest that the initial effect of fenfluramine is mediated via a serotonergic mechanism while the delayed, but long-lasting suppression of plasma renin activity, is mediated via a noradrenergic mechanism.

Progesterone stimulates sexual behaviour in female rats by increasing 5-HT activity on 5-HT 2 receptors

We have investigated the possibility that there is a correlation between female sexual receptivity and CNS serotonergic activity. In ovariectomized rats primed with a submaximal steroid regime of 2 micrograms oestradiol benzoate (OB) plus 0.2 mg progesterone (P), so that only a proportion were receptive, the 5-hydroxyindoleacetic acid:5-hydroxytryptamine (5-HT) ratio in the hypothalamus was greater in the receptive compared to the non-receptive rats. When rats were primed with 10 micrograms OB alone, there was no difference in 5-HT activity in the receptive and non-receptive animals. Depletion of hypothalamic 5-HT levels by p-chlorophenylalanine methyl ester (PCPA) inhibited behaviour normally induced by OB plus P and this could be reversed by 5-hydroxytryptophan. PCPA had no effect in animals made receptive by OB alone. This indicates that P, but not OB, exerts its stimulatory effect on sexual behaviour via increasing 5-HT activity. Administration of 5-HT into the lateral, but not 3rd ventricle stimulated sexual behaviour and systemic injection of the selective 5-HT1 and 5-HT2 agonists, Ru 24969 and MK 212, inhibited and stimulated behaviour, respectively. It is suggested that 5-HT has a dual effect on female sexual receptivity acting via different systems, the inhibitory tract acting on 5-HT1 and the stimulatory tract on 5-HT2 receptors.

A possible mechanism of the tyramine-induced head-twitch response

The effects of monoaminergic drugs on the head-twitch response (HTR) induced by intracerebroventricular (i.c.v.) injection of tyramine (TyA) in mice pretreated with safrazine, a monoamine oxidase inhibitor, were compared with effects on the response induced by the i.c.v. injection of serotonin (5-HT) in safrazine-pretreated mice. The HTR induced by both TyA and 5-HT were suppressed by i.p. injection of p-chlorophenylalanine. Chlorimipramine enhanced the 5-HT response but not the TyA response. Dimethothiazine, a serotonergic blocker, reduced both the TyA and 5-HT responses. The i.c.v. injection of p-chlorophenylalanine methylester resulted in a reduction of the TyA response but not of the 5-HT response. The i.c.v. injection of 5,6-dihydroxytryptamine (5,6-DHT) 1 day before the test suppressed the TyA response but enhanced the 5-HT response. The i.c.v. injection of noradrenaline reduced both the TyA and 5-HT responses. The i.c.v. injection of 6-hydroxydopamine, and i.p. injection of α-methyl-p-tyrosine and tolazoline accelerated the TyA response. These results suggest that the TyAresponse may be based on the release of endogenous 5-HT and can be suppressed by the catecholaminergic system.

Actions of pharmacological agents on 5-hydroxytryptamine and dopamine in the cockroach nervous system ( Periplaneta americana L.)

1. 1. Liquid chromatography with electrochemical detection was used to determine concentrations of dopamine, N-acetyldopamine, 5-hydroxytryptamine, and N-acetyl-5-hydroxytryptamine in cerebral ganglion of the American cockroach, Periplaneta americana. 2. 2. Neuroactive drugs were injected into the cockroaches and their effects on biogenic amine levels in cerebral ganglion assessed. Drugs tested included α-methyltyrosine, p-chlorophenylalanine methyl ester, D( + )-amphetamine, reserpine, pargyline, harmaline, and the formamidine pesticides chlordimeform, N-demethylchlordimeform, and N, N-didemethylchlordimeform. 3. 3. Although N-acetyldopamine was easily demonstrable in P. americana cerebral ganglion, N-acetyl-5-hydroxytryptamine could not be detected in cerebral ganglion from normal animals. However, when cerebral ganglia were incubated with 5-hydroxytryptophan or 5-hydroxytryptamine in vitro, a peak with the low oxidation potential and retention time of N-acetyl-5-hydroxytryptamine appeared. 4. 4. p-Chlorophenylalanine methyl ester and α-methyltyrosine, which have dramatic effects on the levels of 5-hydroxytryptamine and catecholamines respectively in mammalian brain, failed to significantly affect the levels of 5-hydroxytryptamine and dopamine in cerebral ganglia from cockroaches pretreated with the drugs. 5. 5. Two possible products of monoamine oxidase activity, dihydroxyphenylacetic acid and 5-hydroxy-indoleacetic acid, were not detected in cerebral ganglion from P. americana. 6. 6. Results of the present study do not support the hypotheses that either MAO inhibition or N-acetyl-transferase inhibition are important modes of insecticidal action of the formamidine pesticides.

Morphine and ethanol as discriminative stimuli: absence of antagonism by p-chlorophenylalanine methyl ester, cinanserin, or BC-105.

It is well established that the effects of morphine and ethanol can function as discriminative stimuli in the rat. The present investigation sought to determine whether discriminated responding mediated by morphine and ethanol is affected by (a) p-chlorophenylalanine methyl ester (p-CPA) an inhibitor of the synthesis of 5-hydroxytryptamine (5-HT) and (b) antagonists of 5-HT. The blocking agents employed were cinanserin, in ethanol-trained subjects, and BC-105, in morphine-trained subjects. No evidence of antagonism was obtained with either BC-105 or cinanserin. Results with p-CPA were somewhat variable but no consistent or statistically significant reduction in the efficacy of morphine or ethanol as discriminative stimuli was observed. The present results with p-CPA are at variance with earlier work by Schechter (1973) and by Rosecrans et al. (1973) who observed antagonism of the stimulus effects of ethanol and morphine, respectively, following depletion of 5-HT with p-chlorophenylalanine.

Alterations in the behavioral effects of LSD by motivational and neurohumoral variables

Forty naive male albino rats were trained to press a bar on a fixed-ratio (FR 32) schedule of water reinforcement. They were then divided into two groups, one of which (N = 20) recieved 5 min of extra water 12 hr before each experimental session; the other group N = 20) received no extra water. Half of the animals in each group was then given three daily doses (100 mg/kg) of the tryptophan hydroxylase inhibitor p-chlorophenylalanine methyl ester (PCPA) while the remaining animals were given control injections of the PCPA vehicle. Ten days following the last administration of PCPA (or vehicle) all animals were given a low dose of LSD (20 μg/kg). Bar-pressing behavior was significantly disrupted only in those animals receiving both PCPA and extra water. Central (whole brain) concentrations of serotonin (5-HT) were significantly lower in all animals which had been treated with PCPA. These results, along with those previously reported, suggest that amount of deprivation can be an important determinant of both the ability of drugs to alter behavior and the dependence of such alterations upon underlying neuronal activity.

Prolonged effects of p-chlorophenylalanine on the blood pressure of conscious normotensive and DOCA/saline hypertensive rats.

1. In deoxycorticosterone acetate (DOCA) saline hypertensive rats a single dose of p-chlorophenylalanine methylester (PCPAME) (400 mg/kg i.p.) produced a significant fall in blood pressure (20-43 mmHg) which lasted for at least 8 days and was accompanied by a parallel depletion of brain stem 5-hydroxytryptamine (5-ht) but not of noradrenaline (NA). 2. In normotensive rats single doses of PCPAME (200 and 400 mg/kg i.p.) produced a significant hypotension (15-20 mmHg) after a latent period of 5 days. An initial pressor response (12 mmHg) was observed at the higher dose level only on day 3. 3. The hypotensive response to PCPAME (200 mg/kg i.p.) in normotensive rats was not modified by pretreatment with 5,6-dihydroxytryptamine (5,6-DHT; 50 mug i.c.v.) or 6-hydroxydopa (6-ohda; 3 X 250 mug intracerebroventricularly). 4. It is concluded that the hypotensive response to PCPAME in normotensive rats in independent of brain stem depletion of 5-HT and is probably not mediated by the formation of a false transmitter substance acting via central noradrenergic inhibitory pathways. The mechanism involved in the antihypertensive response to PCPAME in DOVA/saline hypertensive rats has yet to be defined.

P-Chlorophenylalanine-Induced Alterations in the Behavioral Effects of 5-Hydroxytryptophan

Pretreatment of rats with p-chlorophenylalanine methyl ester (PCPA-ME) causes a normally non-effective dose of 5-hydroxytryptophan (5-HTP) to disrupt bar-pressing at times when serotonin (5-HT) concentration is depleted. There does not appear to be any correlation between the initial, behaviorally-distruptive effects of PCPA-ME and the action of this compound on 5-HT biosynthesis.

P-Chlorophenylalanine-induced alterations in the behavioral effects of 5-hydroxytryptophan.

Pretreatment of rats with p-chlorophenylalanine methyl ester (PCPA-ME) causes a normally non-effective dose of 5-hydroxytryptophan (5-HTP) to disrupt bar-pressing at times when serotonin (5-HT) concentration is depleted. There does not appear to be any correlation between the initial, behaviorally-distruptive effects of PCPA-ME and the action of this compound on 5-HT biosynthesis.

Female lordosis behavior in estrogen-primed male rats treated with p-chlorophenylalanine or α-methyl- p-tyrosine

The injection of a-methyl- p-tyrosine methylester hydrochloride, 100 mg/kg ip, facilitated lordosis in castrated male rats, treated daily with 50 μg/kg estradiol benzoate. Injection of p-chlorophenylalanine methylester hydrochloride, 150 mg/kg ip, did not affect the lordosis behavior. The effect of a-methyl- p-tyrosine was abolished by the administration of l-3,4-dihydroxyphenylalanine methylester hydrochloride, 50 mg/kg ip, in combination with an inhibitor of peripheral dopa decarboxylase, N 1 -( dl-seryl)- N 2 -(2,3,4-trihydroxybenzyl) hydrazine, 50 mg/kg ip. The results suggest that catecholamine synthesis inhibition is responsible for the facilitation of lordosis behavior seen in esttogen-pretreated male rats after administration of central monoamine-depleting agents. The mechanism whereby drugs that interfere with central monoamine neurotransmission facilitate lordosis behavior in rats is discussed.

Mating behavior in the male rat treated with p-chlorophenylalanine methyl ester alone and in combination with pargyline.

The methyl-ester-hydrochloride of p-chlorophenylalanine alone (50 or 150 mg/kg i.p.) and in combination (4×100 mg/kg i.p.) with pargyline (100 mg/kg i.p.) caused a shortening of the ejaculation latencies in male rats and an increase in the number of intromissions per minute. No changes were observed in other components of the sexual behavior including intromission latency, post-ejaculatory interval and the number of mounts and intromissions preceding ejaculation.

P-chlorophenylalanine methyl ester: an aphrodisiac?

P-Chlorophenylalanine methyl ester and the ester plus pargyline have been reported to facilitate sexual mounting behavior in animals, but these studies have shown only a facilitation of homosexual mounting. The present study indicates that these agents do not enhance the probability or frequency of heterosexual interactions in rats.

Kinetic and fluorescent histochemical analysis of the serotonin compartments in rat pineal gland

Kinetic and fluorescent histochemical changes of rat pineal serotonin (5HT) were found after injecting p-chlorophenylalanine methyl ester (PCP), desipramine (DMI), or reserpine, alone or in combination. These changes indicate that the 5HT stores in sympathetic nerves amount to about 26 mμg/pineal which is approximately 30% of the total pineal store. Since this 5HT store almost completely disappears after DMI, a drug which blocks amine transport in neurons, it is concluded that 5HT is synthesized in pinealocytes and is subsequently taken into axons. The turnover rate of 5HT in the pinealocytes was estimated to be about 100 mμg/pineal/hr. The half-life of decline of 5HT stored in axons after PCP treatment is about ten times faster than that of pinealocytes.