The effect of PEA on serotonin and catecholamines

BIOCHEMICAL

Vol. 103, No. 3,1981 December

AND

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15, 1981

COMMUNICATIONS Pages 1107-1111

THE EFFECT OF PEA ON SEROTONIN AND CATECHOLAMINES Martin

M. Widelitz'

and Charles

P. Hable,

II2

1-2 VA Medical Center, Coatesville, PA 19320 and 1 Departments of Psychiatry and Human Behavior and Pharmacology Jefferson Medical College Thomas Jefferson University Philadelphia, PA 19107, USA

Received

October

X,1981 Summary

Differences of behavior in rats have amine or 6-phenylethylamine (PEA). polyribosomes. It would appear that ior and polyribosomal disaggregation of dopamine, while PEA acts through through norepinephrine and dopamine.

been noted when using d-amphetBoth these drugs can disaggregate amphetamine affects both behavthrough the release and activity serotonin and to a lesser degree

It has been demonstrated that the naturally-occurring sympathomimetic amine B-phenylethylamine (PEA) can induce a behavioral state in rats which consists of sniffing, licking of the cage and horizontal or circular movements of the head which is known as head checking. This is similar to the stereotypic behavior pattern found with the use of amphetamine. There are differences however, that have already been described (1). These consist of a Straub tail (a rigid plasticity of the tail often raised perpendicular to the long axis of the animal), abduction of the hind quarters and a tremor-like state. It has been suggested that amphetamine, an indirect acting agonist, stimulates release of dopamine from the presynaptic terminals and this, acting on the postsynaptic receptor, in turn is responsible for the behavioral events as well as the disaggregation of polyribosomes. Trulson and Jacobs (2) and Zemlan (3) described a behavior pattern which consisted of a Straub tail, tremor, abduction of hind limbs, rigidity, reciprocal forepaw treading and lateral head weaving which was attributed to acThese investigators utilized ptivity at central serotonin mediated synapses. It has also been shown that chloroamphetamine to induce the serotonin release. both amphetamine and PEA can induce polyribosomal disaggregation and inhibit protein synthesis (4, 5, 6, 7). Our interest in the present work evolves from previous work which strongly suggested that when utilizing amphetamine, dopamine is the agonist which induces polyribosomal disaggregation in the animals that have the classical stereotypic pattern (sniffing, licking, wire biting, head checking). In this investigation we wished to determine the mode of action of PEA. Methods Fifty-four CD rats (Charles River) weighing between 250-350 mg were individually housed at room temperature and were maintained on a 12:12 hour (7 a.m. 7 p.m.) light/dark schedule with Purina laboratory chow and water continuously available. The animals were then divided into four groups, those receiving saline acting as the control group, then injected with 100 mglkg of PEA. A second group of animals receiving methysergide 4 mg/kg, a serotonin blocking agent, was then injected with PEA 100 mg/kg 90 minutes later. A third group given pimozide 25 mglkg, which inhibits the action of dopamine on postsynaptic recep-

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tors, was then injected with PEA 100 mg/kg 30 minutes later while the last group given diethyldithiocarbamate (DDC) 400 mgfkg, which blocks the production of All chemnorepinephrine, was then injected with PEA 100 mg/kg 90 minutes later. After observing the behavior patterns icals were given as single IP injection. for 30 minutes the rats were decapitated. The brains were rapidly removed and homogenized in buffer containing 25 mM Tris-HCl, (pi-l 7.5), 5 mM Mg (CH3C00)2, 5 mM Z-mercaptoethanol, 100 mM KC1 and 10% sucrose (w/v). Polysomes were pelleted through 2 M sucrose as previously described (4), suspended in buffer containing 25 mM Tris-HCl (pH 7.5), 5 mM Mg (CH3COO)2, 100 mM KC1 and 5 mM 2-mercaptoethanol and utilized for velocity sedimentation in sucrose gradients. One to 2 ml polysome samples containing 6.0-12.0 A 260 units were layered on 10-35X linear sucrose gradients of 26 ml volume containing 25 mM Tris-HCl (pH 7.5), 100 mM KC1 and 3 mM MgC12. Centrifugation was carried out in an SW 25 rotor of a Beckman L2-50 ultracentrifuge at 76,000 x g x 2-l/2 hours (3-5' C). Gradients were fractionated using a model UA5 Isco density gradient fractionator and maintained at 254 nm. Peak areas of subunits (40 S and 60 S) and monomers (80 S) and that of the polymeric region (> SO S) on the optical density tracings were measured and the percentage calculated. Results All 17 rats given saline prior to PEA exhibited the specific abControl: normal behavior pattern consisting of a straubed tail and abduction of the hind Other less specific characteristics seen as well with amphetamine, quarters. such as sniffing and head checking, were also observed in these same animals., In this group, 100% of the animals showed significant disaggregation of the polyribosomes (polyribosomes are considered intact when there is greater than 68% absorbance in the region greater than 110 S). Pimozide: When pimozide was utilized before the injection of PEA, 67% of the animals exhibited a behavioral syndrome consisting of a straubed tail, abduction of the hind quarters and tremor. 25% of the animals in this group showed the sniffing, licking or head checking, typical of amphetamine induced stereotypic behavior. DDC: 100% of the group treated patternconsisting of Straub tail group 42% also showed polyribosomal

with DDC prior to PEA manifested and abduction of hind quarters. disaggregation.

a behavior In this

Methysergide: In this group 69% of the animals showed a straubed tail and abduction of hind quarters when methysergide was used as a serotonin blocking agent. In these animals only 23% showed polysomal disaggregation. The methysergide treatment group had the highest mean percentage of intact polyribosomes (71.5%) and was the only group whose mean percentage of polysomes fell within the range of intact polysomes (greater than 68% absorbance in the region greater than 110 S). The mean values and standard error of the polymeric particles are shown in Table 1. Statistical Analyses I Non-Parametric: Comparison of treatment groups to control group. Comparison of the number of animals showing disaggregation of polysomes as opposed to the number of animals showing intact polysomes. The difference in the proportion of rats having disaggregated polysomes versus intact PolYsomes between the methysergide treatment group and the PEA control group was very highly significant (x2=16.31, P<.QOl). The difference somes versus intact

in the proportion polysomes between

of rats having disaggregated brain polythe DDC treatment group and the PEA con-

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Effects

of

Treatment

8lOCHEMlCAL

Phenylethylamine

on Brain

w/kg

AND

Polyribosomes

BIOPHYSICAL

TABLE and

1 Behavior

% Exhibiting Specific Syndrome

No. Rats Tested

RESEARCH

Following

Injection

Mean 2 Polysomes

PEA

COMMUNICATIONS

of

Psychotropic

Drugs

% Rats Having Dissociated Polysomes

+ SE

4

13

69

71.46

1.74

23

25

12

67

65.31

1.94

67

DDC-PEA

400

12

100

67.02

2.79

42

Saline-PEA

---

17

100

37.95

2.06

100

Methysergide-PEA Pimozide-PEA

trol groups was significant (x2=5.89, P<.O5), the difference in the methysergide group.

although

not

as significant

The difference in the proportion of rats having disaggregated somes versus intact polysomes between the Pimozide treatment group control group was not significant (x2=4.07, Pi.05). Statistical Analyses II Parametric Analysis: Comparison control. In comparing polyribosomes in control and treatment that the analysis of variance revealed an overall very highly ference (F=7.50, df 3,50, P<.OOl). Dunnet's T test for differences between treatments a highly significant difference between the methysergide the control group (T=4.39, P<.OOl). group, Pc.05). control

The DDC treatment was significantly

group, while different

The Pimozide group however, group (T=2.34, Pc.05).

as

brain polyand the PEA

of all means with group, we found significant dif-

and controls revealed treatment group and

not as significant as the methysergide from the saline control group (T=2.95,

was not

significantly

different

from

the saline

Discussion In this investigation we attempted to find the mechanism of action of PEA in affecting behavior and the disaggregation of polyribosomes. In utilizing PEA we observed a behavior syndrome that was significantly different from that found with amphetamine and as already reported, it was thought this behavior pattern might be due to serotonin activation of appropriate receptors (1). To avoid confusion because of a significant overlap between the animals behavior when given amphetamine or PEA we have isolated two signs which in the acutely treated animals appear to be specific for PEA and not found in amphetamine treated animals. These are the Straub tail and the abduction of the hind quarters of the animals. Utilizing an intraperitoneal injection of 64 mg/kg of PEA along with pargyline, and MAO inhibitor, Moja et al. (8) have reported a stereotypic behavior pattern. This has also been the case with Borison and Diamond (9), who gave 50 mg/kg of PEA daily for 21 days. In analyzing and rating the animal's abnormal behavior pattern however, neither group describes the specific behavior differences that we have found with the single injection of PEA as noted above. The difference in behavior that we described with PEA is unlikely dose related, as with doses of 50 mg/kg we have found these specific manifestations in a significant number of rats, while these two specific events were not reported by

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Moja et al. (8) injecting 64 mg/kg of PEA along with paragyline ing to explain the reason for the difference in behavior between wondered if the technique of chronic injection of PEA that was and Diamond (9) might be important. The possibility of strain another area that might have played a role in this phenomenon. 50 mg/kg of PEA in both Wistar and Sprague-Dawley rats however, tered the specific behavior pattern in a consistent fashion.

COMMUNICATIONS In attemptIP. our groups, we used by Borison specificity was When we injected we again encoun-

In the present work we attempted to find the chemical ligand responsible for the disaggregation of polyribosomes after the injection of PEA. It has been established (10) that serotonin has the ability to disaggregate polyribosomes. When we utilized methysergide as a serotonin blocker only 23% of the rats had polyribosomal dissociation as opposed to higher percentages in the other cateThe difference was highly significant when compared with gories (see Table 1). the percentage of polysome dissociation in the saline control and the other treatment groups. As alluded to previously (l), it was thought that PEA might stimulate both dopamine and serotonin release. With the present data however, dopamine does not appear to play a significant role in polysome disaggregation, while norepiBornephrine appears to be more significant but not as important as serotonin. ison and Diamond (9) have suggested a role for norepinephrine in PEA induced "stereotypies" as they state that DDC (a dopamine B-hydroxylase inhibitor) in selectively antagonizes PEA stereoa dose that depletes brain norepinephrine, typic behavior. These investigators (9) utilized 20 mglkg of DDC while we used 400 mg/kg. They found approximately 60% of their rats manifested stereotypic behavior, while all of our animals showed the abnormal specific behavior pattern. Several facts may explain the quantitative difference in the behavior observed between the two groups. Borison and Diamond (9) utilized 50 mg/kg of PEA while we used 100 mg/kg. They treated their rats daily for approximately 21 intraperitoneal injection. When DOPA, an amino days, while we used a single acid precursor of norepinephrine, as well as dopamine is given in a dose that increases brain norepinephrine both PEA and amphetamine stereotypies are potentiated. Borison and Diamond (9) have also stated that noradrenergic blockade antagonizes the chronic effects of PEA administration and that in chronic PEA induced stereoOur typic behavior both noradrenergic and dopaminergic pathways are involved. work would suggest however, that while norepinephrine may play a role in polyribosomal disaggregation after the use of PEA, the major factor in the process Sloviter (11) wished to determine if of disaggregation appears to be serotonin. PEA shares a common serotonergic property with amphetamine which he says produces serotonergic effects in relatively large doses. In his investigation he found that PEA (80 mg/kg) had no significant effect on whole brain 5 hydroxytryptamine (5-HT) or dopamine concentration 10 minutes after injection, a time coincidental with display of the syndrome. He interprets these results as suggesting that the acute behavior after PEA was not mediated by a large general increase in brain 5-HT. Unlike our findings he found amphetamine twice as potent as PEA in evoking the behavioral response described above, although there Sloviter goes on to say that p-chlorophenylais no mention of a Straub tail. lanine (pCPA) a tryptophan hydroxylase inhibitor decreased norepinephrine, dopamine and serotonin to 75, 68 and 12% of control respectively. Animals given pCPA did not prevent the 5-HT behavioral syndrome caused by PEA but did prevent the syndrome caused by amphetamine. The prevention by pCPA of the amphetamine 5-HT syndrome was reversed by an injection of 5 hydroxytryptophan (5-HTP) in treated rats. His conclusions are that PEA possesses serotonergic activity in vivo and that this property is also possessed by amphetamine. While PEA andamphetamine are structurally similar the behavior they stimulate in rats is by different mechanisms. Amphetamine appears to work by releasing serotonin from depots while PEA is independent of endogenous 5-HT levels and acts directly on 5-HT receptors.

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Promoting the thesis that the behavioral syndrome of Straub tail, abduction of hind quarters and polyribosomal disaggregation are related, one is disturbed by the apparent disparity in the methysergide group. Here we see that 60% of the animals had an abnormal behavior pattern, described above with PEA, while only 23% of the animals manifested dissociated polyribosomes. The possibility exists that PEA may disaggregate polyribosomes in specific brain areas, If such as the limbic system to a greater extent than we see in whole brain. this were the case, it might explain the apparent inconsistency already noted where the effect on behavior is greater than the effect in aggregation of polyribosomes in whole brain. One might also consider an alternative possibility that the behavior pattern may depend on the relative amounts of serotonin, dopamaine and norepinephrine (the balance hypothesis). If in fact this is the case, the inconsistency between aberrated behavior and polysome dissociation might be related to the different sensitivities of these processes to the relative amounts of these fractions. As already noted however, it would appear that under the conditions of these experiments, serotonin is the major factor affecting polysome aggregation and behavior following injection of PEA. The inhibition of serotonin activity by methysergide in relation to dopamine and norepinephrine might have a greater effect on the polyribosomes than on behavior. Acknowledgements This research was supported Acknowledgement and appreciation the manuscript.

by US Veterans is due to Mrs.

Administration, Washington, E. Theresa Snyder for typing

DC.

References 1.

2. 3. 4. 5. 6. 7. 8. 9. 10. 11.

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