A demonstration that 5-hydroxytryptamine administered peripherally can affect sexual behavior in male rats

In the male rat, subcutaneous injections for 7 days of 20 mg/Kg B.W./day of 5-hydroxytryptamine creatinin sulphate (5-HT), caused remarkable i$\text{n}$hibitory effects on sexual behavior.The mount and intromission latencies were incre$\text{a}$sed in rats treated with 5-HT, whereas ejacul$\text{a}$tion latency in the few rats treated with 5-HT that it achieved, was similar to that obtained in control rats.The mount and intromission frequencies were decreased in the rats treated with 5-HT.The mean inter-intromission interval (MII) and post-ejaculatory interval were prolonged in rats treated with 5-HT.These data provide evidence for the role of peripheral 5-HT in regulating sexual behavior of - male rats.     

Deamination of 5-methoxytryptamine, serotonin and phenylethylamine by rat MAO in vitro and in vivo.

Pretreatment of rats with clorgyline, a selective inhibitor of MAO-A, significantly inhibited the $\text{in vivo}$ deamination of intraventricularly administered serotonin (5-HT) and 5-methoxytryptamine (5-MT), but not phenylethylamine (PEA). Pretreatment with d, l-deprenyl, a selective inhibitor of MAO-B, significantly inhibited the $\text{in vivo}$ deamination of all three substrates. Brain and liver homogenates from rats pretreated with clorgyline showed a decreased ability to deaminate ($\text{in vitro}$) 5-MT and 5-HT, but not PEA. Homogenates from animals pretreated with d,l-deprenyl showed a decreased capacity to deaminate PEA, but not 5-MT or 5-HT. Clorgyline, when added to brain and liver homogenates, selectively blocked the deamination of 5-MT and 5-HT, but not PEA, whereas, d,l-deprenyl blocked the deamination of PEA without affecting that of 5-MT or 5-HT. In addition, 5-MT was found to be 100 X more potent than PEA at inhibiting the $\text{in vitro}$ deamination of 5-HT. These findings suggest that 5-MT and 5-HT are favored substrates for MAO-A $\text{in vitro}$ and $\text{in vivo}$. However, $\text{in vivo}$, significant amounts of 5-MT and 5-HT can also be deaminated by MAO-B.     

Opioid properties of beta-lipotropin fragment 60-65, H-Arg-Try-Gly-Gly-Phe-Met-OH.

The pharmacologic activity of the hexapeptide fragment corresponding to the amino acid fragment 60–65 in β-lipotropin, (β-LPH-(60–65)) was studied $\text{in vitro}$ and $\text{in vivo}$. In binding assays on synaptosomal plasma membrane the peptide was found to be equipotent to met-enkephalin, but behaved differently to cations; in contrast to met-enkephalin both Mn⁺² and Na⁺ enhanced the binding of β-LPH-(60–65) to synaptosomal plasma membrane. On both the quinea pig ileum and mouse vas deferens β-LPH-(60–65) inhibited contractions elicited by electrical stimulation and each effect was reversible by naloxone. On the guinea pig ileum β-LPH-(60–65) was equipotent to met-enkephalin and 0.5 as potent as normorphine but on the vas deferens it was 4.6 times more potent than normorphine. The activities of β-LPH-(60–65) appear to be due to the intact compound rather than to its conversion to met-enkephalin, since the peptide extracted from the ileum assay was found to behave identically as β-LPH-(60–65) with high pressure liquid chromatography. When β-LPH-(60–65) was administered centrally to mice and rats, no overt central actions were observed and an antinociceptive effect could not be demonstrated. Nor did β-LPH-(60–65) antagonize morphine action or precipitate the withdrawal syndrome in morphine dependent animals. It is concluded that the good agreement which generally exists between $\text{in vitro}$ and $\text{in vivo}$ assay procedures for opiate-like activity of morphine and its surrogates does not necessarily hold for the endogenous peptides with similar actions.     

Characterization of rat pulmonary monoamine oxidase.

The substrate specificities and kinetics of rat lung monoamine oxidase (MAO) have been studied. Utilizing the irreversible MAO inhibitors, clorgyline and deprenyl, rat lung was shown to possess at least two types of MAO, A and B. Tyramine was a substrate for both forms of the enzyme, whereas 5-hydroxytryptamine (5-HT) was a preferred substrate for the A-form. In contrast to most other tissues, 2-phenylethylamine was not solely a B-type substrate for the rat lung MAO and some metabolism by the A-type was apparent $\text{(}\text{B}\text{A}\text{=}\text{80}\text{20}\text{)}$. Using tyramine as substrate the ratio A/B was shown to be $\text{55}\text{45}$. Rat pulmonary MAO-B was inhibited by deprenyl and the kinetics of MAO-A studied. The Km values for the A-form for tyramine, phenylethylamine and 5-HT were relatively similar and were 270, 244 and 170 μM respectively. Whereas, when the A-form was inhibited by clorgyline, the Km values for the B-form were found to differ considerably: 330, 42 and 850 μM for tyramine, phenylethylamine and 5-HT respectively.     

Effects of 4-methyl-α-ethyl-tyramine, 4, α-dimethyl-tyramine and tetrabenazine on the release and uptake of 5-hydroxytryptamine by human blood platelets invitro

4-Methyl-α-ethyl-tyramine and its 4, α-dimethyl analogue release 5-HT from human blood platelets $\text{in}$$\text{vitro}$. At lower concentrations they inhibit the uptake of 5-HT into platelets. Tricyclic antidepressant drugs do not block 5-HT release by these compounds. On removal of the depletor, platelets recover their ability to take up 5-HT; platelets preloaded with exogenous 5-HT lose the same proportion of amine as those containing only endogenous 5-HT. Tetrabenazine behaves similarly, but its actions are partial, whereas those of the tyramines are more complete. The temperature dependence of spontaneous and drug-induced 5-HT release has been measured. The results are discussed in terms of the action of these drugs and with special reference to the use of human blood platelet as a model of a 5-HT-containing nerve ending.     

Acetylated and nonacetylated forms of beta-endorphin in rat brain and pituitary

Extracts of rat posterior intermediate pituitary and extracts of brains from normal and hypophysectomized rats were separated by gel filtration chromatography and fractions were analyzed by both a classical β-endorphin radioimmunoassay and by a radioimmunoassay specific for α-N-acetyl β-endorphin. In posterior intermediate pituitary extracts, more than 90 percent of the β-endorphin-sized immunoreactive material was α-N-acetylated. In extracts of brains from normal rats, less than 2 percent of the β-endorphin-sized immunoreactive material corresponded to α-N-acetylβ-endorphin, whereas in brains from hypophysectomized animals, no α-N-acetylβ-endorphin-like material could be detected. Immunofluorescence on normal brain sections, using either affinity purified antibodies to α-N-acetylβ-endorphin or conventional β-endorphin antibodies, showed no α-N-acetylβ-endorphin immunoreactivity in β-endorphin neurons. Only in brain sections which had been acetylated $\text{in}\text{vitro}$ prior to immunostaining could α-N-acetylβ-endorphin-like material be detected in the β-endorphin neurons. These results suggest that—in contrast to the cells in the intermediate lobe of the pituitary—the β-endorphin in brain neurons is not α-N-acetylated and that the small amount of α-N-acetyl β-endorphin which can be found in extracts of brains from normal animals is probably of pituitary origin.     

Microdetermination of norepinephrine, 3,4-dihydroxyphenylethylamine, and 5-hydroxyyptamine from single extracts of specific rat brain areas

The conditions reported by Toru and Aprison (4) for extracting ACh in specific brain areas were tested to determine whether 5-HT, NE, and dopamine were also extracted quantitatively. It was found that the extraction solution used in brain ACh determinations, 15% 1N formic acid plus 85% acetone ($\text{v}\text{v}$), was also excellent for extraction of NE, 5-HT, and dopamine from different brain areas. Experimental conditions are given for the microdetermination of all three biogenic amines in such a single extract of a specific rat brain area. The methods are based on previously published fluorometric methods; these have been scaled down or modified slightly to permit analyses of small aliquots. The concentration of 5-HT, NE, and dopamine in the telencephalon, diencephalon plus mesencephalon, pons plus medulla oblongata, and cerebellum of the rat are also reported using the described micromethods after extraction with 15% 1 N formic acid plus 85% acetone ($\text{v}\text{v}$).     

Altered lipoxygenase metabolism and decreased glutathione peroxidase activity in platelets from selenium-deficient rats

Washed platelets from selenium-deficient and control rats were incubated with [1-¹⁴C]-arachidonic acid and the lipoxygenase and cyclooxygenase products were identified by gas chromatography/mass spectrometry. Platelets from selenium-deficient rats showed a three to four-fold increased synthesis of the lipoxygenase-derived isomeric trihydroxy fatty acids, 8,9,12-trihydroxy-5,10,14-eicosatrienoic acid and 8,11,12-trihydroxy-5,9,14-eicosatrienoic acid. A major reduction in glutathione peroxidase activity was also observed in platelets from deficient rats. These results support the interpretation that these trihydroxy fatty acids arise from breakdown of the primary platelet lipoxygenase product $\text{L}$-12-hydroperoxy-5,8,10,14-eicosatetraenoic acid (12-HPETE) under conditions in which its reduction to the $\text{L}$-12-hydroxy product (12-HETE) by a selenium-dependent glutathione peroxidase is limited. Further-more, these results indicate a specific function for selenium in platelet metabolism of essential fatty acids.     

5α-reduction of an anti-androgen TSAA-291, 16β-ethyl-17β-hydroxy-4-estren-3-one,by nuclear 5α-reductase in rat prostates

Inhibition of 5α-reduction of testosterone by an anti-androgen TSAA-291 (16β-ethyl-17β-hydroxy-4-estren-3-one) was studied in rat ventral prostates and the metabolic conversion of ³H-TSAA-291 was examined both $\text{in vitro}$ and $\text{in vivo}$. In the $\text{in vitro}$ experiment using nuclear 5α-reductase of the prostate, 5α-dihydrotestosterone formation from ³H-testosterone was inhibited in a competitive manner by the anti-androgen. In the $\text{in vitro}$ experiment using ³H-TSAA-291, 5α-reduction of the anti-androgen occurred. One, 2 and 4 hr after an intravenous administration of 140 μCi/rat of ³H-TSAA-291 to castrated rats, the unchanged TSAA-291 accumulated in higher amounts in the ventral prostate than in the plasma, skeletal muscle and levator ani muscle, thereby indicating the selective uptake of the anti-androgen by the androgen target organ. No appreciable amounts of the 5α-reduced metabolite of TSAA-291 were detected in the prostate, thus suggesting that TSAA-291 itself may be responsible for the anti-androgenic properties. The inhibitory potency on the 5α-reductase activity of several other 16β-substituted androstane and estrane analogues was also examined.     

Confirmation of an unexpected brain O-methylation pattern for the dopamine-derived tetrahydroisoquinoline,salsolinol

Using high resolution capillary gas chromatography, we have unequivocally separated two possible (6- and 7-)mono-O-methylated tetrahydroisoquinoline metabolites in rat brain after acute intraventricular administration of salsolinol, a cyclized dopamine/acetaldehyde derivative. 7-O-Methylsalsolinol (salsoline) constituted 94–98% of the two isomers in five brain regions examined. These results confirm the report by Bail $\text{et}$$\text{al}$. that salsolinol is largely O-methylated $\text{in}$$\text{vivo}$ (presumably by brain catechol-O-methyl transferase) on the hydroxyl situated “para” in the parent dopamine molecule. In comparison, dopamine itself, administered intraventricularly to pargyline-pretreated rats, was O-methylated exclusively on the “meta” hydroxyl group.     

Opiate receptor affinities and behavioral effects of enkephalin: Structure-activity relationship of ten synthetic peptide analogues

Synthetic met- and leu-enkephalin bind to rat brain opiate receptors with $\text{1}\text{2}$ and $\text{1}\text{7}$ the affinity of morphine. The aromatic hydroxyl moiety of the tyrosine residue is critical for receptor binding. Intracranial microinjection of met-enkephalin requires very high doses to produce an evanescent, naloxone reversible analgesia and stuperous immobility, presumably because of its rapid enzymatic degradation. Leu-enkephalin fails to elicit analgesia.     

Roles of 5-hydroxytryptamine and dopamine as neurotransmitters eliciting release of erythrophorotropic hormones in the fiddler crab,Uca pugilator

This article reviews the endocrinological, pharmacological and biochemical evidence ascribing neurotransmitter roles for 5-hydroxytryptamine (5-HT, serotonin) in eliciting the release of red pigment-dispersing hormone (RPDH) and for dopamine (DA) in stimulating the release of red pigment-concentrating hormone (RPCH) in the fiddler crab, $\text{Uca pugilator}$. 5-HT produces red pigment dispersion in intact crabs, but only indirectly. Likewise, DA evokes red pigment concentration $\text{in vivo}$ but it has no effect on red chromatophores (erythrophores) in isolated legs. The data obtained with 5-HT and DA agonists and antagonists on red pigment translocation $\text{in vivo}$ and $\text{in vitro}$, are consistent with their neurotransmitter candidacies in evoking the release of these erythrophorotropic hormones.     

Temperature-sensitive high affinity [3H]serotonin binding: Characterization and effects of antidepressant treatment

Characterization of temperature-sensitive [³H]serotonin (5-HT) binding sites (1 and 4 nM Kd sites) revealed complex inhibition by neuroleptics and serotonin antagonists. There was no simple correlation with affinities for S₁ and S₂ receptors. $\text{In vivo}$ pretreatment (48 h before) with mianserin did not alter Bmax or Kd for the 1 nM Kd [³H]5-HT site, although [³H]ketanserin (S₂) densities were decreased by 50%. This suggested that possible S₂ components of [³H]5-HT binding must be negligeable, even though ketanserin competed with high affinity (IC₅₀ = 3 nM) for a portion of the 1 nM Kd [³H]5-HT site. Low concentrations of mianserin inhibited the 1 nM Kd [³H]5-HT site in a non-competitive manner, as shown by a decrease in Bmax with no change in Kd after $\text{in vitro}$ incubation. The complex inhibition data may therefore represent indirect interactions through another site.     

Regulation of cyclic AMP in heart and gill of Aplysia by the putative neurotransmitters dopamine and serotonin.

Serotonin (5-HT) and dopamine (DA), but not several other putative neurotransmitters, stimulate cyclic adenosine-3',5'-monophosphate (cAMP) production in slices of $\text{Aplysia}$ gill. Furthermore, 5-HT but not DA increases cAMP in slices of the heart of $\text{Aplysia}$. Several lines of evidence indicate that the receptors are distinct entities; however, no drugs were found to block one receptor without affecting the other.     

Effect of synthetic estrogen on platelet aggregation and vascular release of PGI2-like material in the rabbit

Treatment of adult female New Zealand white rabbits with ethinyl estradiol, the synthetic estrogen used in many oral contraceptives, results in a significant increase in $\text{in vivo}$ aggregation. This alteration in platelet behavior is accompanied by diminished vascular release of antiaggregatory PGI₂ (prostacyclin)-like material. Addition of a progestin prevents the change in platelet aggregation seen with the estrogen alone. Diminished vascular PGI₂ release may be an important factor in the pathogenesis of thrombotic occurrences experienced by some oral contraceptive users. $\text{In vivo}$ platelet aggregation may be of value in identifying individuals at risk of developing thrombotic disturbances while taking oral contraceptives.     

The metabolism of dopamine in rat caudate nucleus can be increased by in vivo “dialysis” perfusion cannulae

The development of dialysis cannula techniques coupled with high performance liquid chromatography and electrochemical detection (HPLC-EC) has provided a means to continuously sample extracellular fluid from deep brain structures $\text{in vivo}$. The present studies show that with HPLC-EC analysis of the acid metabolites of dopamine (DA) and 5-hydroxytryptamine (5-HT) in samples from dialysis cannulae implanted in the caudate nucleus of anaesthetized rats, it is possible to determine the time course of the response of dopamine- and 5-HT containing neurones to administration of monoamine oxidase inhibitors and haloperidol. The tissue concentrations of the DA and 5-HT metabolites were also determined at the conclusion of each individual experiment in both the caudate nucleus containing a cannula and in that without a cannula. In perfusion experiments where no drug was administered the content of the DA metabolites, but not that of the 5-HT metabolite, were significantly elevated in the caudate nucleus containing the cannula as compared with the contralateral tissue. These increases occurred whether the cannula was perfused or not, suggesting that the presence of the cannula may have been causing a lesion which altered the activity of the DA neurones. These results emphasize the importance of tissue analysis in conjunction with the dialysis experiments, especially where perfusion sample contents of the monoamine metabolites are being measured as a reflection of the effects of behavioural manipulation or drug treatment on endogenous neuronal activity.     

Low platelet monoamine oxidase activity and chronic marijuana use

Mean platelet monoamine oxidase (MAO) activity in 26 consecutively-studied male marijuana smokers was significantly lower than in a comparable group of non-marijuana smoking males. In addition, the level of current marijuana use reported by the subjects was significantly and inversely correlated with MAO activity. No acute reduction in MAO activity was found in response to smoking a marijuana cigarette containing 15 mg of delta-9-THC. Significant $\text{in vitro}$ inhibition of MAO activity by THC was detected only at THC concentrations above 10^?5M, approximately 100 times the peak plasma concentrations seen $\text{in vivo}$ following smoking.     

Inhibitor of pyrimidine metabolism from tumor tissues

Inhibitors of normal rat liver 5′-nucleotidase and dUMP kinase $\text{in vitro}$ were found in rapidly proliferating tissues, such as Yoshida sarcoma. Two inhibitors were separated from Yoshida sarcoma by zone electrophoresis, gel filtration on Sephadex G-200 and DEAE-cellulose column chromatography. One inhibited both 5′-nucleotidase and dUMP kinase, while the other inhibited only dUMP kinase. These inhibitors were not detectable in normal rat liver. They were induced in regenerating rat liver and present in rapidly proliferating tissues, such as Yoshida sarcoma and Ehrlich ascites tumor and rat marrow cells. These inhibitors were heat labile. One had a large molecular weight (500,000>) and the other a small molecular weight ($\text{Ca}\text{.}$ 50,000).     

Sodium-dependent transport of 5-hydroxytryptamine (serotonin) by canine blood platelets

The transport of 5-hydroxytryptamine (5-HT) was shown to be strongly dependent on the presence of Na⁺ in the incubation medium whereas divalent cations were without effect. The K_m for the Na⁺ requirement was 16.8 mm. The addition of Na⁺ to Na⁺-depleted platelets restored maximum 5-HT transport within 3 min. The affinity of the 5-HT carrier for its substrate was directly proportional to the concentration of Na⁺; however, below 25 mm Na⁺ unique reversible morphological changes in platelet shape occurred as revealed by scanning electron microscopy which resulted in a drastically reduced affinity for 5-HT. K⁺, choline (Ch⁺), or Li⁺ could be used as counterbalancing cations to maintain osmolarity, and the affinity for 5-HT was also dependent on the concentrations of these ions. Ouabain as well as various ionophores at low concentrations inhibited 5-HT uptake. The inhibition was the result of the destruction of the $\text{Na}{}^{\mathrm{+}}\text{K}{}^{\mathrm{+}}$ gradient across the cytoplasmic membrane. Ionophores, however, did not cause the depletion of either intracellular ATP or 5-HT.