Brain Research Bulletin, Vol. 15, pp. 183-189, 1985. a Ankho International Inc. Printed in the U.S.A. 0361-9230185 $3.00 + .OO

Behavioral and Neurochemical Effects of Deprenyl and p-Phenylethylamine

in Wistar Rats

A. J. GREENSHAW, A. V. JUORIO AND A. A. BOULTON

Psychiatric Research Division, Saskatchewan Health, Cancer and Medical Research Building University of Saskatchewan, Saskatoon, Saskatchewan

Received 13 November 1984

GREENSHAW, A. J., A. V. JUORIO AND A. A. BOULTON. Behavioral and neurochemicaf eficts of deprenyl and Pphenylerhylnmine in Wistar rats. BRAIN RES BULL 15(2) 183-189, 198X-The effects of I-deprenyl(l-16 mg kg-‘, 3.5 hr) on brain levels of endogenous /3-phenylethylamine were assessed in animals under three conditions: (1) experience of lateral hypothalamic self-stimulation; (2) electrode implantation but no self-stimulation experience; (3) no surgical inter- vention. The increase in striatal levels of /3-phenylethylamine with I-deprenyl treatment was attenuated in the self- stimulation condition relative to conditions (2) and (3). This differential effect of I-deprenyl was not observed at the level of the hypothalamus. Administration of I-deprenyl did not affect self-stimulation behavior. Equivalent analysis of P-phenylethylamine levels was carried out using animals injected with P-phenylethylamine (0.5-4 mg kg-‘, 0.5 hr 1P and I-deprenyl(4 mg kg-‘, 3.5 hr SC). Injected /3-phenylethylamine with deprenyl pretreatment increased self-stimulation rates; concomitant striatal levels of approximately 190 ng g-’ of @phenylethylamine were observed and were associated with increased brainstem S-HIAA but no change in striatal HVA, indicating possible involvement of 5-HT in this response to P-phenylethylamine. It is proposed that experience of electrical hypothalamic stimulation may alter endogenous striatal /3-phenylethylamine metabolism, possibly via an alteration of mechanisms governing synthesis and/or catabolism.

P-Phenylethylamine I-Deprenyl Intracranial self-stimulation Reinforcement Lateral hypothalamus Striatum Homovanillic acid 5-hydroxyindoleacetic acid Monoamine oxidase Endogenous brain amines

TO a large extent research directed to an analysis of aminer- gic systems which may regulate behavior has focussed on the catecholamines and 5-hydroxytryptamine [ 191. Nevertheless there has been increasing interest in the possible functional roles of other endogenous aryl alkyl amines [4]. One such compound, /I-phenylethylamine, has received considerable recent attention [5,16]. This amine occurs naturally in mam- malian brain, highest levels being observed in the corpus striatum and the hypothalamus [ 111. The heterogeneous dis- tribution, subcellular localisation and rapid turnover of this amine indicate a probable physiological role in neural regu- lation [3, 10, 111. This possibility is strengthened by the ob- servation that iontophoretically applied /3-phenylethylamine may specifically potentiate the effects on unit activity of concurrent applications of dopamine and norepinephrine Pll.

The /3-phenylethylamine moiety forms the basic carbon structure of amines which have sympathomimetic properties [2] and the structural and functional similarity between P-phenylethylamine and amphetamine (CX methylphenyl- ethylamine) has led to the proposal that /3-phenylethylamine may in fact act as an endogenous amphetamine [33]. Abnor- mal levels of endogenous /3-phenylethylamine and its metabolites are claimed to be involved in the aetiology of a number of psychiatric disorders [5, 33, 401 and there is evi- dence which suggests a role for endogenous P-phenylethylamine in neural substrates of reward or rein- forcement [ 161. Nevertheless, there is a paucity of data relat-

ing levels of endogenous or exogenous P-phenylethylamine to behavioral activity under controlled laboratory condi- tions. *

The observation that behavioral responses may be main- tained by electrical stimulation of local brain areas (i.e., in- tracranial self-stimulation) has yielded a useful paradigm with which to investigate neurochemical correlates of rein- forcement [28]. Within this area few studies have examined the possible role of P-phenylethylamine. Administration of this amine to rats pretreated with the monoamine oxidase inhibitor (MAOI) iproniazid results in a reward-specific enhancement of self-stimulation [35]. The contention that endogenous /3-phenylethylamine may be related to rein- forcement has also received some support from studies of the self-administration of this compound by laboratory animals [32,34] and by the observation that P-phenylethylamine may be effective in the conditioned place preference paradigm [ 151.

If P-phenylethylamine metabolism in the central nervous system is related to reinforcement processes, then levels of endogenous P-phenylethylamine may be affected by expo- sure to a reinforcing stimulus. A corollary of this hypothesis is that changes in @phenylethylamine levels will give rise to changes in reinforced behavior. These related hypotheses have been examined fairly extensively with regard to cate- cholamine metabolism in the brain [37]. Nevertheless little attention has so far been paid to brain P-phenylethylamine metabolism in this respect.

183

184 GREENSHAW, JUORIO AND BOULTON

The effects of lateral hypothalamic stimulation and of ~pheta~e on rat brain ~-p~nyle~yl~ine levels have previously been investigated [8]. In this study there were no measured effects of the experimental variables on brain /3-phenylethylamine levels. It is apparent, however, that this lack of an alteration in /3-phenylethylamine levels may be related to the extremely rapid turnover of this amine [lo]. Although it is possible that measurement of the principal metabolite of this amine (phenylacetic acid) may reveal stim~tion-educed changes in ~-phenylethyl~ine levels, it is evident that a significant amount of phenylacetic acid may be derived from another source, phenylpyruvic acid, possibly as a direct consequence of phenylalanine catabolism [38,24]. Adopting an alternative strategy, it was hypothesised that a reinforcement-induced change in brain ~-phenyle~yl~~e levels may be revealed by using an MAO1 and comparing the relative increase in levels of this amine between animals with self-stimulation experience, electrode implantation, and without surgical intervention re- spectively.

In considering a possible physiological role for ~-phenylethyl~ne in this context, a further aim of the present study was to assess the extent to which levels of this amine are associated with changes in self-stimulation behav- ior. Thus, we have also assessed the inlluence of MAO inhi- bition in combination with a range of doses of exogenous p-phenylethylamine on self-stimulation behavior in relation to the measurement of brain levels of this amine.

I-Deprenyl was chosen as an appropriate MAO inhibitor for this study as, at low doses, this compound selectively inhibits the type B form of MAO which is the enzyme which governs the oxidative deamination of p-phenylethyiamine in viva 1391.

METHOD

Maie Wistar rats (Charles River Canada, Montreal) weighing 180-200 g were used. The animals were individu- ally housed on a 12 hr light-dark cycle at a temperature of 20+ 1°C. Food and water were freely available.

Animals were randomly assigned to one of three major test conditions: unoperated (IJ), implantation with hypotha- lamic electrodes (0), and i~l~~~on with hy~~~~ic electrodes with training for intracranial self-stimulation (S) respectively. All animals were subjected to a routine of daily handling for a period of approximately two to five minutes. Control animals were not exposed to the behavioral test apparatus.

Animals allocated to electrode treatment groups were anaesthetised with Nembutal (40-50 mg kg-‘) and, using a Kopf stereotaxic instrument, each rat was implanted unilat- erally with a twisted bipolar stainless steel electrode (Plastic Products Ltd., Roanoke, VA, MS 30311) directed to the me- dial forebrain bundle at the level of the lateral hypothalamus [26]. The coordinates (mm from bregma) were AP + 1.5, lat- eral +1.5, ventral -8.5. The electrodes were secured to the skull with stainless steel screws and dental acrylic.

After a seven day recovery period animals allocated to the self-stimulation condition were trained to lever-press on a fted-interval3 second schedule of electrical hy~~~~ic stimulation. On this schedule, as described previously [ 171,

animals were allowed to self-regulate the duration of each train of stimulation. When stimulation was available a lever-press served to initiate a train of stimulation which remained on until the animal released the lever. The animals were trained in standard test chambers (Coulbourn Insts.), each equipped with house-light and a light above the re- sponse lever which was switched on during the delivery of stimulation, Hypothalamic stimulation was provided from a constant current source (36-106 @A rms, 60 Hz sine-wave) which was connected to the rats through gold track slip-rings (Lehigh Valley). Throughout the test sessions current inten- sity for each subject was monitored on an oscilloscope as the voltage drop on a 10k R resistor connected in series with the rat. CBM 2000 microcomputers served to control the exper- iments and to record behavioral responses online. The over- all rate of lever pressing, the self-regulated duration of hypo- thalamic stimulation, and the duration of responses occur- ring in the absence of stimulation (i.e., during the fixed inter- val) served as behavioral measures. Test sessions were of 30 minutes duration. After 5 to 7 training sessions on the self- stimulation schedule each animal was randomly allocated to a drug treatment condition.

Six groups of self-stimulators were used to assess the behavioral effects of I-deprenyl (0, 1, 2,4, 8, 19 mg kg I). A further four groups of self-stimulators were used to investi- gate the behavioral effects of @-phenylethylamine (0.5. 1 .O, 2.0, 4.0 mg kg-.‘) in combination with I-deprenyi (4.0 mg kg-‘). I-Deprenyl was injected subcut~eously 3.5 hr prior to behavioral testing. p-Phenylethylamine was injected intra- peritoneally immediately prior to behavioral testing. The in- jections were counterbalanced so that animals assigned to the I-deprenyl condition without p-phenylethylamine were injected immediately prior to behavioral testing with the ve- hicle. The present low doses oft-phenyIethylamine were not assessed in the absence of I-deprenyl in this study as previ- ous experiments in our laboratory have failed to detect ef- fects on self-stimulation of p-phenylethylamine at less than 25 mg kg‘-’ [J8].

Drags

The drugs used were I-deprenyl which was kindly supplied by Professor J. Knoll, and @-phenylethylamine hydrochlo- ride obtained from Sigma Chemical Company, St. Louis, MO. I-Deprenyl and /3_phenylethylamine HCl were each dis- solved in 0.9% saline and injected in a volume of 1 ml kg ‘. Vehicle injections were administered in an equivalent vol- ume.

Nrurochemical Analysis

Immediately following completion of behavioral testing the self-stimulators were killed by cervical dislocation. Their brains were rapidly removed and the striata and hypothala- mus of each subject were dissected out over ice. These samples were then placed into a container with dry ice and stored for not more than 2 hr at -20°C.

/3-Phrnylethylamine Estimution

The striata or the hypothalami of two rats were weighed and homogenised in 1 ml of 0.1 N HCI containing ethylene diamine tetraacetic acid disodium salt (EDTA, 1 mg mt--‘) and ascorbic acid (5 mg ml-‘) and 50 ng of tetradeutero

/3-PHENYLETHYLAMINE AND REINFORCEMENT 185

TABLE 1

EFFECTS OF I-DEPRENYL ON ENDOGENOUS BRAIN &PHENYLETHYLAMINE LEVELS IN ICSS, OPERATED AND UNOPERATED CONTROL RATS

I-deprenyi mg kg-’ 0 1 2 4 8 16

Striatum

Hypothalamus

Striatum

Hypothalamus

Striatum

Unoperated Controls (U)

*;:: @)

,::: (5)

Operated Controls (0)

2;:: (Q

k::; (@

ICSS (S)

2;:: 0)

2;:; (4)

2;:: (3)

::I. (7)

*::; (7)

::I$’ (4)

2;:: (4)

:;I$’ (6)

+-:I: (4)

16.9”’ k4.5 (5)

9.4 22. I (4)

17.4” c5.6 (6)

18.6”‘““’ (6) r 16.0

13.7” ~2.8 (4)

7.7 22.5 (3)

19.%“$ (7) +3.6

19.9”‘““’ (7) 2 17.0

14.48 A4.0 (5)

15.9 *14.2 (5)

Effects of a range of doses of I-deprenyl (SC 3.5 hr) on striatal and hypothalamic levels of endogenous /?-phenylethylamine in each of the U/O/S treatment conditions. Values are means ? S.E.M. (number of determinations in parentheses) in ng g-j of fresh tissue. Comparisons were made with the Newman-Keuls test (a=O.O5) following analysis of variance.

Superscripts denote: ;-different from 0 mg kg-‘; “-different from 1 mg kg-‘; ‘-different from 2 mg kg-‘; “-different from 4 mg kg -‘; ‘---different from 8 mg kg -I.

phenylethylamine as an internal standard. The amines in the tissue homogenate were derivatised with 5-di- methylamino-1-naphthaiene suiphonyl (dansyl) chloride and the resultant derivatives were extracted into toluene- ethylacetate (91 v/v), evaporated to a small volume and separated chromatographically in two different unidimen- sional systems 1221. /3-Phenylethylamine was estimated by a high resolution mass spectrometric selected ion monito~ng technique using deuterated @phenylethylamine as an inter- nal standard. Blanks of 0.1 N HCI to which were added 50 ng of deuterated @-phenylethylamine were carried throughout the procedure. These gave blank values of 200 pg and thus enabled quantitation of as little as 200 pg of p- phenylethylamine in the tissue samples. Complete details concerning this procedure have been described [l I].

For comparison with each of these self-stimulation groups equivalent neuro~hemic~ analyses were performed on un- operated control animals and on control animals impianted with a hypothalamic electrode but untested for self- stimulation. These groups were included to permit an analysis of the relative influences of electrode implantation and of behavioral test experience on the neuro~hemical vari- ables. It should be noted that under these conditions each group of animals was killed 4 hr after injection with I-deprenyl or vehicle and 0.5 hr after injection with ~-phenylethylam~e or vehicle respectively.

The injection times were based, for deprenyl, on previous

studies of MAO inhibition [30] and, for /3_phenylethylamine, on pilot testing in our laboratory.

To determine the relative influence of these drugs in dopamine and S-hydroxytryptamine (5HT) activity, levels of homovanillic acid (HVA) and 5hydroxyindoleacetic acid (S-HIAA) were assessed in the striata and brainstem of un- operated control rats.

5-Hydroxyindole Acetic Acid Determinations

These determinations were carried out in the brain stem (including the colliculi, medulla oblongata and pons to the obex). The tissues from two rats were pooled for each de- termination. The tissues were homogenised in 0.1 N NC1 containing ascorbic acid (5 mg ml--‘), deproteinised, extra- cted into butylacetate and determined fluorimetrically [14]. Checks on recoveries of 100 ng of added 5-hydroxy~ndole acetic acid were carried out in each experiment; the percent- age of recovery was 75.5r1.4 [15]: mean*SEM, number of experiments in parentheses; and the results were corrected accordingly.

Homovanilk Acid Estimations

The striata of two rats were homogenised in 0.1 N hydro- chloric acid, extracted with butylacetate and then into tris buffer (0.05 M) and estimated fluo~met~ca~y [I]. The per-

186 G~EENSHAW, JUGRIG AND BOU~T(~N

TABLE 2 STRIATAL LEVELS OF INJECTED #%PHENYLETHYLAMINE IN ICSS OPERATED AND UNOPERATED

CONTROL RATS AFTER I-DEPRJZNYL PRETREATMENT

0

Injected ~-Phenylethyiamine mg kg ’

0.5 1.0 2.0 4.0

Unoperated CORtrOk (u)

:;I; (6) z;:f (4) $:y (6) 174.5” (6) 464 3;’

zY30.5 * (4) t 131.5

Operated Controls (0)

118.6’ (8) +23.6 ,“;I: (8)

ICSS (S) +!::; (4) 4;:; (4)

186 8;’ . (4) e82.4

Effects of a range of doses of exogenous /.%phenyiethyiamine (IP OS hr) in combination with Ldeprenyl(4 mg kg-’ SC 3.5 hr) on striatal levels of P-phenylethylamine in each of the U/O/S treatment conditions.

Values are means + S.E.M. (num~r of dete~inations in parentheses) in ng g”’ of fresh tissue. Compari- sons were made with the Newman-Keuls test (a=0.05) following analysis of variance.

Superscripts denote: “-different from 0 mg kg-‘; “-different from 0.5 mg kg-‘: ‘-different from 1 mg kg-‘; d-diEerent from 2 mg kg-‘; “-different from 4 mg kg- 1.

centage recovery of 200 ng of added HVA was 76.61t7.1 [S], mean+-SEM; the results were corrected accordingly.

These determinations were carried out for 4 mg kg-’ 1-deprenyl in the presence and absence of 2 and 4 mg kg-’ of p-phenylethylamine with respect to HVA; and for 4 mg kg-’ I-deprenyl in the presence and absence of 0.5, 1.0, 2.0, 4.0 mg kg-’ of ~-phenylethyl~ine with respect to S-HIAA.

RESULTS

Behavioral Analysis

For each of the behavioral measures each group’s per- formance was expressed as the mean percentage of that for the daily session immediately prior to drug treatment. These dam were subjected to analysis of variance. Administration of 1-deprenyl 3.5 hr prior to testing did not result in any significant changes in response rate, F(5,46)=0.82, p>O.O5; self-regulated duration of electrical stimulation, F(5,46)=0.06, p>O.OS; or the duration of non-reinforced responses, F(5,46)=0.99, p>O.O5. Administration of /3- phenylethyl~ine to animals pretreated with I-deprenyl resulted in a significant enhancement of response rates, F(4,39)=7.50, p<O.OOl, but failed to affect either the self- regulated duration of electrical stimulation, F(4,39)= 1.09, p>O.5, or the duration of non-reinforced responses, F(4,39)= 1.89, p>O.2. Newman-Keuls multiple comparisons (ar=O.O5) revealed that the rate effect was due to a significant increase in this measure with 4.0 mg kg’ of @-phenylethylamine (199.1*26.3% with deprenyl 4 mg kg-’ plus 4 mg kg-’ ~-phenylethyl~ine relative to 101.4+6.% for deprenyl 4 mg kg-’ and 101.4+6.% for saline treated controls; values are mean+s.e.).

Neurochetnical Analysis

The effects of administration of I-deprenyl on endogenous levels of @phenylethylamine are displayed in Table 1 for both striatal and hypothalamic samples.

Analysis of variance conducted with the striatal samples revealed both a significant increase in endogenous @-phenylethylamine levels in response to administration of I-deprenyl, F(5,76)=28.74, p<O.OOl, and between unoper- ated, operated and self-stimulation (U/O/S) treatments a

difference in endogenous P-phenylethylamine levels, F(2,76)=5.53, p<O.Ot. The interaction between U/O/S treatments and effects of l-deprenyl was not statistically sig- nificant, F( 10,76)=0.97,p>O.5. From the data in Table 1 it is evident that I-deprenyl markedly increased endogenous levels of fi-phenylethylamine in the striatum to between 12-and Hi-fold of control levels with each group at the high- est dose. Notably, the maximal increases were observed with the unoperated and the operated control groups relative to the groups of self stimulators. Multiple comparisons (Newman-Keuls, (r=O.O5) revealed that the dose-dependent nature of this response to I-deprenyl was greatest in the un- operated groups and least in the groups of self-stimulators. With each of the U/O/S treatments each dose of I-deprenyl resulted in a significant elevation of &phenylethylamine. With the unoperated controls there was a si~i~c~t differ- ence in p-phenylethylamine levels between the 16 and 8 mg kg-’ groups relative to 2 mg kg-’ and between the 16 mg kg--’ group and the 4 mg kg-’ groups. With the operated controls the only further significant comparison was between the 16 mg kg-’ and the 1 mg kg-’ group. The setf-stimulators, how- ever, exhibited no dose-dependent response to I-deprenyl as only the sahne vs. I-deprenyl group comparisons were signif- icant.

Analysis of variance conducted with the hypothalamus samples revealed a significant increase in P-phenyl- ethylamine in response to administration of I-deprenyl, F(5,68)=6.11, p<O.OI. There was, however, no signi- ficant effect of the U/O/S factor, F(2,68)=0.27, p>OS, and no significant interaction, F(10,68}=0.49, p>O.5. From Table 1 it is evident that for hypothalamic samples, as with the striatum, I-deprenyl markedly increased endogenous levels of p-phenylethylamine. With the hypothalamic sam- ples this increase was up to between 4- and S-fold with each group at the highest dose. Multiple comparisons (Newman- Keuls, tu=O.O5) revealed that the dose-response to I-deprenyi was significant between 16 mg kg-i and all other treatments for the unoperated control groups. The operated controls exhibited increases at 16 mg kg-’ and 8 mg kg- ’ relative to ail other treatments. With the self-stimulators, however, equivalent tests between the saline and I-deprenyl groups did not reveal any signiftcant individual comparisons.

p-PHENYLETHYLAMINE AND REINFORCEMENT 187

TABLE 3

EFFECTS OF l-DEPRENYL AND OF p-PHENYLETHYLAMINE ON STRIATAL HVA AND BRAINSTEM 5-HIAA LEVELS IN UNOPERATED RATS

mg kg-’

Striatal HVA Levels

Dose of Injected P-Phenylethylamine

0 2.0 4.0

l-deprenyl 0 p;; (5)

4.0 .“; (3

I:; (4)

:;‘4 (4)

mg kg-’

Brainstem 5-HIAA Levels

Dose of Injected P-Phenylethylamine

0 0.5 1.0 2.0 4.0

I-deprenyl

The effects on striatal HVA and brainstem 5-HIAA levels of a range of doses of exogenous P-phenylethylamine (IP0.5 hr) alone or in combination with I-deprenyl(4 mg kg-l SC 3.5 hr).

Values are means & S.E.M. (number of determinations in parentheses) in ng g-’ of fresh tissue. With respect to HVA, administration of I-deprenyl resulted in an overall decrease in this measure; the effects of P-phenylethylamine were not significant. 5-HIAA was signifi- cantly decreased by I-deprenyl. P-Phenylethylamine significantly increased 5-HIAA levels. See text for further details.

As the analyses described above revealed statistically significant differences between the U/O/S treatment groups for striatal but not hypothalamic samples, a post-hoc analysis of the relative response to deprenyl between these regions was conducted for each level of the U/O/S grouping factor.

In accord with the data presented above, these analyses revealed an overall significant difference between the striatal and hypothalamic fi-phenylethylamine levels for the un- operated controls, F( 1,39)=7.97, ~~0.01, and for the oper- ated control groups, F(1,66)=5.48, ~~0.05. There was, how- ever no significant regional effect with the self-stimulators, F(1,39)=2.85, p>O.lO.

The striatal levels of P-phenylethylamine observed in animals pretreated with 4 mg kg-’ I-deprenyl and injected with P-phenylethylamine or the saline vehicle are displayed in Table 2. The control groups (i.e., 0 mg kg-l P-phenylethylamine plus 4.0 mg kg-’ I-deprenyl) are the same groups of animals as represented in the I-deprenyl dose-response analysis of Table 1. With this analysis, maxi- mal P-phenylethylamine levels were observed at the highest injected dose of this amine. The degree of elevation was >38-fold in unoperated controls; >22-fold in operated con- trols; and > 16-fold in self-stimulators.

Analysis of variance with these data revealed a significant increase in striatal P-phenylethylamine, F(4,71)=35.26, p<O.OOl, but, despite consistently lower mean values in the self-stimulation groups, no significant effect of the U/O/S treatment factor, F(2,71)= 1.90, p>O. 1. Multiple compari- sons (Newman-Keuls, a=0.05) revealed that for each treat- ment group the elevation of /3-phenylethylamine in all animals injected with this amine was significantly greater than that achieved with I-deprenyl alone. Due to the increas-

ing variability in levels of this amine with increasing injected doses, no other individual between-dose comparisons reached statistical significance.

The effects on striatal HVA and brainstem 5-HIAA of I-deprenyl at 4.0 mg kg-’ in combination with injected doses of P-phenylethylamine are displayed in Table 3. Analysis of variance conducted on these data revealed, for HVA: a sig- nificant effect of I-deprenyl, F(1,22)=21.49,p<0.001, but no effect of injected p-phenylethylamine, F(2,22)=0.22, p>O.5, and no significant interaction, F(2,22)= 1.18, p>O.5. This ef- fect of I-deprenyl reflected an overall decrease in HVA in response to this compound (Newman-Keuls test, (r=O.O5). With the brainstem 5-HIAA samples analysis of variance revealed a significant effect of I-deprenyl, F( 1,43)=21.49, p<O.OOl, and of injected p-phenylethylamine, F(4,43)=2.97, ~~0.05. There was, however, no significant interaction, F(4,43)=0.40, pbO.5. From the data in Table 3 it is evident that these effects reflect a decrease in 5-HIAA in response to I-deprenyl and an increase in this compound in response to /3-phenylethylamine. Multiple comparisons (Newman- Keuls, a!=O.O5) did not, however, reveal any significant in- dividual group differences.

DISCUSSION

The present experiments were conducted primarily in order to explore the possibility that brain fl-phenyl- ethylamine metabolism may be affected by experience of electrical self-stimulation of cells in the lateral hypo- thalamus. In view of the rapid turnover of this amine, the MAO1 I-deprenyl was used to inhibit the catabolism of en- dogenous /3_phenylethylamine.

With the differential increase in striatal /3-phenyl- ethylamine in response to I-deprenyl alone, two possibilities

188

are evident: (1) Electrode implantation and self-stimulation experience may have altered the rate of synthesis of j3-phenylethylamine and therefore the availability of this substrate for MAO. (2) MAO activity may have been altered in response to electrode implantation and self-stimulation experience.

The former possibility may be plausible in relation to the shared biosynthetic pathways for catecholamines and P-phenylethylamine [51. This effect could be metabolically related to an increase in striatal dopamine activity, as fore- brain catecholamine turnover is enhanced by electrical self-stimulation at medial forebrain bundle sites [ 13,361. With respect to a possible alteration in enzyme activity, an in- crease in type B monoamine oxidase activity would result in a shift to the right in the dose-response curve to deprenyl. Notably, however, due to the high rate of turnover of /3-phenylethylamine under normal conditions this effect would not necessarily result in a change in basal levels of this amine. This is consistent with the present fmdings. Never- theless such a change in enzyme activity may, theoretically, also result in an increased rate of catabolism of an injected load of the exogenous amine. This tendency was evident in the present study in response to 1 to 4 mg kg-’ of p-phenylethylamine, i.e., lower mean levels were observed in the self-stimulators. This effect was not, however, statisti- cally significant. As /3-phenylethylamine is lipophilic [27], such an effect may have been obscured by diffusion of the amine between different brain regions and, possibly to a greater extent, between the brain and the periphery. It is interesting to note that an induction of striatal type B monoamine oxidase in rat brain has been reported after hemitransection [7]. However, it is evident that within the present dose-range for deprenyl maximal inhibition of MAO-B may be expected. Nevertheless, it is evident that deprenyl is a relatively weak MAO inhibitor in terms of elevating /3-phenylethylamine levels in brain (e.g., contrast the present results with those of [30]). This relative potency remains unexplained and may be related to aspects of drug action other than MAO inhibition. Additional experiments are necessary to investigate further the alternative hypothe- ses outlined above. In any event, the present results indicate that /%phenylethylamine metabolism in the rat striatum may be affected by electrode implantation and by self-stimulation training.

The analysis of HVA and of 5-HIAA levels in unoperated rats revealed that 4 mg kg-’ of I-deprenyl decreased the catabolism of 5-HT and of dopamine. The former result indi- cates that this pretreatment was not restricted to the inhibi- tion of type B monoamine oxidase but also inhibited the type A form of the enzyme to some extent [391. It is notable that the additional (exogenous) p-phenylethylamine treatment in- creased 5-HIAA levels, possibly indicating a release of S-I-IT at the doses tested [12]. HVA levels were unaffected, how- ever, by p-phenylethylamine under these conditions.

The range of doses of I-deprenyl used in this study did not alter the present behavioral measures. This finding stands in contrast to a previous brief report of effects of I-deprenyl on self-stimulation [Z]. It is apparent, however, that I-deprenyl is converted to amphetamine in vivo [23, 29, 311 and the time-course of this effect may have contributed to the re-

GREENSHAW, JUORIO AND BOULTON

ported stimulant actions of deprenyl [25]. In the present study a pre-treatment to test interval of 3.5 hr was chosen. There is effective inhibition of MAO under these conditions, however, amphetamine levels after deprenyl are very low in brain at this time [29].

As administration of I-deprenyl did not result in any changes in the present behavioral measures a range of intra- peritoneally injected doses of /3-phenylethylamine were as- sessed in animals pretreated with I-deprenyl. This experi- ment yielded a significant increase in overall rates of re- sponding at the highest dose of P-phenylethylamine. This effect is consistent with the results of an earlier study [35] which demonstrated rate-enhancing effects of this amine in self-stimulating rats pretreated with the MAOI iproniazid. Under the present conditions, a minimum sttiatal level of 187 ng g-’ of /3-phenylethylamine accompanied a behavioral re- sponse to this amine. This represents a very large increase in striatal concentrations of this amine and the relevance of this effect in other than pharmacological terms is therefore ques- tionable. Nevertheless it must be emphasized that endoge- nous supplies of @phenylethylamine are normally subject to rapid turnover [lo]. In relation to the neurochemical basis for the present rate-enhancing effects of /3_phenylethylamine, the present lack of effect of this amine on HVA levels is notable. There is some controversy over the contention that the behavioral effects of /?-phenylethylamine may be mediated by dopamine systems [6,20]. The present observa- tion of increased 5-HIAA levels in response to p-phenylethylamine provides supportive evidence for a possible involvement of 5HT as previously suggested [9,12] although it is difficult to make direct comparisons between these studies and the present results in view of either the use of a different MAO inhibitor [ 121 and/or the use of very large doses (or high concentrations) of P-phenylethylamine [6, 9, 12, 201. Nevertheless, the choice of low p-phenylethylamine doses in combination with MAO inhibition is clearly appro- priate in the present context.

The alteration in endogenous p-phenylethylamine levels in response to electrode placement and self-stimulation experience in the present study may be a reflection of altera- tion in a p-phenylethylamine system involved in neural regu- lation. It is evident that the possibility that electrical hypo- thalamic stimulation per se may have contributed to the present alteration of /3-phenylethylamine metabolism must be considered. As no yoked-control animals (or animals with neutral placements) were included in the present study the question of to what extent the reported effect is related to experience of response-contingent stimulation remains open. Further research is necessary to explore these proposals.

ACKNOWLEDGEMENTS

The authors wish to thank Professor J. Knoll, Department of Pharmacology, Semmelweis University, Budapest, Hungary for a generous gift of I-deprenyl, Dr. D. A. Durden for supervising the mass spectrometric analyses, Dr. B. A. Davis for synthesizing the deuterated P-phenylethylamine, M. Mizuno, G. Wheatley and E. P. Zarycki for expert technical assistance and Saskatchewan Health for continuing financial support. A. J. Greenshaw is supported by a Saskatchewan Health Research Board Fellowship.

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