Judith A. Richter

Acetylcholine and choline levels in post-mortem human brain tissue: Preliminary observations in Alzheimer's disease

Abstract Choline and acetylcholine were measured in necropsy brain tissue (temporal cortex and caudate nucleus) obtained from elderly, mentally normal hospital cases and established cases of Alzheimers disease. ACh levels were as expected, extremely low in all cases; in cases with Alzheimers disease, the ACh level was lower in the temporal cortex but not changed in the caudate compared with normal cases (matched for ages and post-mortem sampling delays). The level of choline in Alzheimers disease was not significantly different from the normal in either brain region. The choline levels in the human material were, however, substantially and significantly lower than those obtained from young adult rat cerebral cortex which was cooled after death according to the post-mortem temperature decline in the human cadaver.

Barbiturates: Their in vivo effects and potential biochemical mechanisms

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Release of glutamate and CGRP from trigeminal ganglion neurons: Role of calcium channels and 5-HT1 receptor signaling

BackgroundThe aberrant release of the neurotransmitters, glutamate and calcitonin-gene related peptide (CGRP), from trigeminal neurons has been implicated in migraine. The voltage-gated P/Q-type calcium channel has a critical role in controlling neurotransmitter release and has been linked to Familial Hemiplegic Migraine. Therefore, we examined the importance of voltage-dependent calcium channels in controlling release of glutamate and CGRP from trigeminal ganglion neurons isolated from male and female rats and grown in culture. Serotonergic pathways are likely involved in migraine, as triptans, a class of 5-HT1 receptor agonists, are effective in the treatment of migraine and their effectiveness may be due to inhibiting neurotransmitter release from trigeminal neurons. We also studied the effect of serotonin receptor activation on release of glutamate and CGRP from trigeminal neurons grown in culture.ResultsP/Q-, N- and L-type channels each mediate a significant fraction of potassium-stimulated release of glutamate and CGRP. We determined that 5-HT significantly inhibits potassium-stimulated release of both glutamate and CGRP. Serotonergic inhibition of both CGRP and glutamate release can be blocked by pertussis toxin and NAS-181, a 5-HT1B/1D antagonist. Stimulated release of CGRP is unaffected by Y-25130, a 5-HT3 antagonist and SB 200646, a 5-HT2B/2C antagonist.ConclusionThese data suggest that release of both glutamate and CGRP from trigeminal neurons is controlled by calcium channels and modulated by 5-HT signaling in a pertussis-toxin dependent manner and probably via 5-HT1 receptor signaling. This is the first characterization of glutamate release from trigeminal neurons grown in culture.

The site of action of naloxone in suppressing food and water intake in rats

Abstract Previous studies have shown that naloxone causes a decrease in food and water intake; however, the site of this action has not been determined. We investigated this problem by giving bilateral injections of 15 μg/rat of naloxone into the lateral ventricles of cannulated, food and water deprived rats. This treatment caused a significant decrease in food intake when compared to saline injected controls. Water intake in naloxone-treated animals did not differ significantly from that of saline-treated controls during the one hour test period. The total dose of naloxone given centrally, 15 μg, did not produce a change in eating or drinking if given peripherally. The findings imply that naloxone exerts its effect on food intake at a central site. A dose-related and significant suppression of water intake was seen after treatment with nalaxone peripherally (1, 3, and 10 mg/kg, i.p.) in rats with either subdiaphragmatic vagotomy (vag) or a sham vagotomy (sham). Although a significant suppression of food intake was seen in the sham rats, no supression of food intake was seeen in the vag rats at any dose of naloxone tested. In rats pretreated with methyl atropine (5 mg/kg, i.p.), naloxone (3 mg/kg, i.p.) was equivalent to saline in that it did not decrease food intake. However, nalaxone did cause a significant decrease in water intake in methylatropine pre-treated rats. These results suggest that the suppression of food intake by naloxone has a central site of action which is mediated by the vagus, and specifically by vagal efferents, since the effect was blocked by methylatropine. The results also suggest that naloxones effect on water intake is mediated by a different mechanism than that involved with food intake.

Effects of pentobarbital on the regulation of acetylcholine content and release in different regions of rat brain

Abstract Pentobarbital administered to rats in rivo increased the levels of acetylcholine (ACh) in the cerebral cortex, striatum, hippocampus and pons-medulla but not in the midbrain or cerebellum. It has been suggested that the increased levels of ACh may be the result of an inhibition of release of the transmitter. However, when pentobarbital was tested in vitro, it inhibited potassium-stimulated ACh release from all regions. Release of ACh and tissue levels of ACh were compared directly in cerebral cortex and midbrain in vitro. Stimulation with 25 and 50 mM KC1 enhanced ACh release from both regions but decreased tissue ACh only in the cerebral cortex. Pentobarbital inhibited potassium-stimulated ACh release from both regions but only increased tissue ACh in the cerebral cortex (stimulated with 25 mM KCl). These results suggest that the regulation of ACh synthesis and release is different in the two brain regions and that pentobarbital can inhibit ACh release without necessarily causing a concomitant rise in ACh tissue levels.

A possible role for tyrosine kinases in the regulation of the neuronal dopamine transporter in mouse striatum

The present investigation was undertaken to test the hypothesis that a reduction in the activity of protein tyrosine kinases would result in an alteration in dopamine transport. Genistein, a broad-spectrum inhibitor of protein tyrosine kinases, inhibited dopamine uptake into mouse striatal homogenates with an IC50 of 18 microM. The inhibition by genistein was rapid, reversible and somewhat selective, in that genistein did not inhibit the uptake of choline or GABA under similar conditions. Kinetic analyses indicated that genistein was a non-competitive inhibitor. Another protein tyrosine kinase inhibitor, tyrphostin 23, also inhibited transport but was significantly less potent than genistein. Tyrphostin 25 and lavendustin A were without major effect on dopamine uptake. In addition, the inactive structural analog of genistein, genistein, had no significant effect on dopamine uptake. The inhibition of dopamine transport by 50 microM genistein was accompanied by a reduction in the level of a 110-kDa band of tyrosine phosphoprotein. It is suggested that protein tyrosine kinases play a role in the cascade of events which ultimately lead to regulation of neuronal dopamine transport.

K–STIMULATED ACETYLCHOLINE RELEASE: INHIBITION BY SEVERAL BARBITURATES AND CHLORAL HYDRATE BUT NOT BY ETHANOL, CHLORDIAZEPOXIDE OR 11‐OH‐Δ9‐TETRAHYDROCANNABINOL

Abstract— Thiamylal, methohexital, secobarbital, amobarbital, pentobarbital, phenobarbital and barbital, and also chloral hydrate inhibited K‐stimuiated acetylcholine (ACh) release from rat midbrain slices in a dose related manner. The concentrations required for 50% inhibition were related to the lipid partition coefficient. None of the barbiturates nor chloral hydrate inhibited unstimulated ACh release. Ethanol, chlordiazepoxide and 11‐OH‐Δ9‐tetrahydrocannabinol were without effect on both unstimulated and stimulated ACh release. The results are discussed in terms of other effects of these drugs on the cholinergic system.

CHARACTERISTICS OF ACETYLCHOLINE RELEASE BY SUPERFUSED SLICES OF RAT BRAIN1

—A superfusion system has been used to examine the effects of choline and the utilization of [3H]choline during resting and potassium‐stimulated release of ACh from rat cerebrum slices. The rate of ACh release from unstimulated tissue, 0·25 nmol/g per min, increased 8‐fold when the concentration of KCl in the superfusing medium was increased from 5 to 50 mm. This rate was not maintained, however, but gradually declined to one‐half the peak rate after approx. 30 min. After an initial washout period, choline was released at a rate of 2·5‐5 nmol/g per min, which was equal to 1‐2 × 10−6m in the superfusate. The addition of 1 × 10−5m‐choline to the superfusing medium was required to maintain the stimulated ACh release at near peak rates for 90 min. When hemicholinium‐3 was added to the 50 mm‐KCl medium, the release of ACh reached a peak as usual but then declined to prestimulation rates.

Dopamine D2 receptors increase in the dorsolateral striatum of weaver mutant mice

Dopamine D2 receptors were studied in homozygous weaver mutant mice (wv/wv), heterozygous littermates (wv/+), and normal mice (+/+). Specific [3H]spiperone binding was significantly higher in the dorsolateral part of the striatum in the weaver mutant mice (wv/wv) than in normal mice (+/+). No significant differences among the 3 genotypes were found in other parts of the striatum or in the nucleus accumbens.

Effects of pentobarbital and Ca2+ on the resting and k+-stimulated release of several endogenous neurotransmitters from rat midbrain slices☆

Abstract The release of endogenous amino acids [γ-aminobutyric acid (GABA), glutamate, aspartate, glycine and alanine] and of 5-hydroxytryptamine (5-HT) and acetylcholine (ACh) from rat midbrain slices was examined under various conditions of superfusion. Depolarization with high K+ stimulated the release of all substances examined, but the K+-stimulated release was Ca2+ dependent only for GABA, glutamate, aspartate, 5-HT and ACh. Pentobarbital, although not substantially affecting resting release, inhibited the K+stimulated release of GABA, glutamate, aspartate and ACh markedly and significantly. The effect of pentobarbital on K+-stimulated 5-HT release was not statistically significant in this series of experiments. These results are consistent with the hypothesis that the barbiturate inhibits stimulated transmitter release by inhibiting Ca2+ influx during depolarization. The tissue content of amino acids and 5-HT decreased considerably from fresh tissue levels after 50 min of perfusion in regular low K+ medium; in contrast, ACh tissue levels increased slightly during this time. K+ stimulation resulted in an increased synthesis particularly of GABA, glutamate, glycine, alanine and ACh, for the amount released into the medium was far more than that lost from the tissue. 5-HT and possibly aspartate, on the other hand, were released into the medium on stimulation largely at the expense of tissue stores, under our experimental conditions.