Biomedical subjects

J R Hepler

Publications and source records attributed to J R Hepler.

43 records · Page 3Linked to original sources

PubMed1985

Regulation of cyclic AMP metabolism by muscarinic cholinergic receptors.

The occurrence of muscarinic cholinergic receptor-mediated activation of phosphodiesterase in 1321N1 cells does not represent an isolated phenomenon, since a similar response to cholinergic stimuli is observed in thyroid slices (45) and WI-38 fibroblasts (1,42). Both muscarinic-receptor-mediated inhibition of adenylate cyclase and activation of phosphodiesterase occur in WI-38 fibroblasts (42). Work currently under way in our laboratory is directed toward determining if a guanine nucleotide regulatory protein is involved in the activation of phosphodiesterase in these cells and whether common or separate populations of muscarinic receptors are coupled to these two mechanisms of cyclic AMP metabolism. The analysis of acute hormonal regulation of phosphodiesterase in intact cells is sufficiently complicated to have previously discouraged investigators from pursuing this question in mammalian tissues. The 1321N1 cell line provides a simple model system in which at least one mechanism of hormonal regulation of phosphodiesterase can be examined. In light of the widespread occurrence of muscarinic-receptor-mediated effects on Ca2+ mobilization, it would not be surprising to find that this mechanism represents an important part of cholinergic action in both the peripheral and central nervous systems. Indeed, this system could provide an important regulatory link between Ca2+ -mediated and cyclic-AMP-mediated events in target cells. The potential importance of such a mechanism also need not be restricted to the muscarinic receptor system, since any neurotransmitter or hormone receptor system coupled to events involved in Ca2+ mobilization might produce phenomena similar to that observed for muscarinic receptors in 1321N1 cells. Our studies emphasize that two mechanisms for regulation of cyclic AMP accumulation by muscarinic cholinergic receptors exist. The data to date suggest that separate receptor subtypes are involved in these mechanisms of cholinergic regulation and provide another biochemical basis whereby the well-known interaction of Ca2+ with the cyclic AMP system can be effected. Thus, identification of the molecular events involved in the regulation of PI turnover and its consequences may be crucial in defining the basis of this aspect of cholinergic action. In addition, more extensive analyses of the phosphodiesterase system using cell-free preparations have the potential of providing clues to the molecular basis of this mechanism.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenosine Diphosphate Ribose↗

PubMed1984

Changes in Ca2+ ion activity within unrestrained rat's hippocampus perfused with alcohol or acetaldehyde.

In the freely moving rat, the kinetics of Ca2+ ion activity were determined at circumscribed sites in the hippocampus, which was perfused with ethanol, tertiary-butyl alcohol or acetaldehyde. Initially, a region in CA1 or other cell field of the dorsal hippocampus was prelabelled by microinjection of 45Ca2+ through a permanently implanted guide tube. Then the tip of a concentric push-pull cannula assembly was lowered through the guide tube to the labelled site, and an isotonic artificial cerebrospinal fluid was repeatedly perfused at a rate of 25 microliter/min. Each perfusion was timed for 5.0 min with a 5.0 min interval between each. Once the washout curve of 45Ca2+ activity had begun to approach its asymptote, ordinarily in the midpoint of a series of perfusions, an isotonic solution of ethanol (188-942 mM), tertiary-butyl alcohol (12-580 mM) or acetaldehyde (10-98 mM) was added to the fourth perfusate. Thereafter, the hippocampal site was again perfused with the normal cerebrospinal fluid for the remainder of the experiment. Although the lowest concentration of ethanol exerted no effect on 45Ca2+ ion activity, an intermediate concentration caused mixed effects in either enhancing or suppressing the efflux into the perfusate of this cation. The highest concentration of ethanol produced in most experiments an initial suppression in Ca2+ ion efflux which was followed frequently by an elevation in the release of 45Ca2+. Similar changes in Ca2+ ion activity were produced by tertiary-butyl alcohol, but the magnitude of its effect was generally less than that of ethanol, suggesting that its effect on brain tissue differs from that of ethanol. Acetaldehyde evoked an intense and concentration-dependent enhancement of Ca2+ ion efflux from the perfused tissue at all of the sites in the hippocampus examined. These results suggest that in the unrestrained rat ethanol could unbind Ca2+ ions from hippocampal membranes or retard their uptake into cells of the hippocampus. The dual excitatory and inhibitory effect of ethanol on Ca2+ ion activity corresponds to the electrophysiological effects of this alcohol and could alter neurotransmitter release from neurons in this subcortical structure. The mechanism of action of acetaldehyde is envisaged to be due to its affinity to membrane sulfhydryl groups which alters protein conformation and thus interferes with both Ca2+ channels and Ca2+ binding properties.

Acetaldehyde↗

PubMed1984

Profile of amino acid synthesis in rat hippocampus during push-pull perfusion of ethanol or morphine.

In the unanesthetized rat, the effect of ethanol or morphine on the new synthesis of six amino acids, gamma-aminobutyrate, glutamate, alanine, aspartate, glycine and glutamine, was determined at circumscribed sites in the hippocampus. In the fasted animal, 5.0 microCi of [U-14C]glucose was microinjected in a volume of 1.0 microliter into a discrete region of the hippocampus through a permanently implanted guide tube. After 10-15 min, the tip of a concentric push-pull cannula assembly was lowered through the guide tube to the labelled site, and an isotonic artificial cerebrospinal fluid was continuously perfused at a rate of 25 microliters/min. Samples of perfusate were collected every 5.0 min in a total volume of 125 microliters. During the fourth perfusion, either ethanol (94 or 471 mM) or morphine (0.1 or 1.0 microgram/microliter) was added to the push-pull perfusate. Amino acids and glucose contained in each sample of perfusate were separated by means of a two-dimensional, thin-layer chromatographic procedure. The results show that the 94 mM dose of ethanol acted only to significantly suppress the new synthesis of alanine. Conversely, the higher 471 mM dose of ethanol significantly increased the new synthesis of both glycine and glutamine from the glucose precursor. The effect of the 0.1 microgram/microliter morphine dose on amino acid synthesis paralleled that of the lower dose of ethanol since it evoked a similar decrease in the synthesis of alanine; however, the higher dose of 1.0 microgram/microliter morphine produced no effect. None of the other amino acids in the hippocampus were affected by either of the CNS depressants. Thus, whereas ethanol seems to exert significant stimulating activity on hippocampal synthesis of glycine and glutamine, morphine and ethanol possess the same inhibitory effect on alanine synthesis in this subcortical structure.

Alanine↗

PubMed1983

New multi-cannula pedestal device for micro-injection of drugs into brain tissue or cerebral ventricle.

A new multiple-cannula-pedestal system for micro-injection of drugs directly into either brain tissue or cerebral ventricle is described. Its features include ease of construction from commonly available materials, no specialized machining required, durability and economy. A special aspect of the cannula system is a protective cap containing a bolt which threads onto a nut fixed within the pedestal base. Since the cap cannot be dislodged, potential damage to indwelling stylets and exposed guide tubes is prevented. Moreover, an aseptic preparation is therefore provided so that test compounds can be infused repeatedly over a prolonged period. Finally, the protective caps are interchangeable and the pedestal base itself can be re-cycled for usage in different animals.

Animals↗

PubMed1983

Evaluation of neurotensin's thermolytic action by ICV infusion with receptor antagonists and a Ca++ chelator.

To investigate the mechanisms responsible for the thermolytic action of neurotensin (NT), cannulae for intracerebroventricular (ICV) injection were implanted stereotaxically in Sprague-Dawley rats. Postoperatively, body temperature in the unrestrained rat was monitored continuously by a colonic thermistor probe. NT in a dose of 1.5-4.5 micrograms or CSF carrier vehicle was infused bilaterally in the cerebral ventricle (ICV) in a volume of 7.5 microliters. With the rats kept at an ambient temperature of 22 degrees C, NT given ICV produced a dose-dependent fall in core temperature of greater than 0.8 degree C. Pre-treatment of the animal's cerebral ventricle with amine receptor antagonists, phentolamine (20.0 micrograms), butaclamol (10.0 micrograms), methysergide (20.0 micrograms) or atropine (25.0 micrograms), all similarly infused ICV, failed to alter the hypothermia induced by NT. However, the calcium chelating agent, EGTA, given ICV in a dose of 4.0-8.0 micrograms blocked the thermolytic effect of NT on body temperature in a concentration-dependent manner. These results suggest that the central thermolytic action of NT is not mediated by catecholamine or other aminergic pathways which are implicated in the central mechanisms of thermoregulation. Rather, the peptide may act on a cellular process involving calcium activity in the hypothalamus, presumably to impair the maintenance of the animal's "set-point" for body temperature.

Animals↗

PubMed1983

Glass-plastic push-pull cannula system for regional perfusion of sites in the brain.

A new non-metallic push-pull cannula device is described for the localized perfusion of brain sites in the rat or other animal. Comprised entirely of glass and plastic materials, the cannula is intended for usage in an experiment in which the presence in tissue of metal tubing for delivery of perfusate is methodologically inappropriate. The main features of the cannula include its light weight, a Delrin pedestal base, a micro-glass guide tube, a polyethylene stylet, plastic spacers for adjustment of depth of perfusion, a protective cap for the chronic maintenance of the preparation, low cost and re-usability. The push-pull cannula is of concentric design and is fashioned from glass capillary and polyethylene tubing which are bonded at their common junctions by an epoxy cement. During a push-pull perfusion a special perfusion cap is used not only to hold the cannula firmly in place but also to protect the glass components from external damage. To further coincide with the metal-free requirements of the preparation, bone screws of nylon are used during the surgical implantation of the pedestal base. A prototype experiment illustrating its use in an unrestrained animal also is presented. The special applications for this cannula system are discussed which include its usage in experiments in either the anesthetized or unanesthetized rat exposed to microwave radiation where the presence of metal in the field would result in unwanted 'hot spots' in the cerebral tissue.

Animals↗

PubMedDate not supplied

Action of anorexigenic peptide injected into the brain: dissociation of effect on body weight and feeding in the rat.

Previous reports on the effect of anorexigenic peptide (AXP) administered systemically in the rodent are inconsistent in terms of the effect of the tri-peptide on food intake and body weight. The purpose of this study was to examine the effect of AXP infused into the brain on these measures. In post-pubescent female rats of the Sprague-Dawley strain, guide cannulae were permanently implanted in the lateral cerebral ventricle for repeated intracerebroventricular (ICV) infusion. Postoperatively, measures of food and water intake and body weight were obtained every day at the same time. After a 7-day base-line period, AXP was infused bilaterally in a total volume of 15 microliter and in a dose of either 0.25 microgram (n=7) or 1.25 micrograms (n=5), with artificial CSF vehicle serving as the control solution (n=6). ICV infusions were given once daily for 20 consecutive days, after which the same intake and body weight measures were recorded for another 7-day period. The rats given 0.25 micrograms AXP showed a significant suppression in weight gain with the overall slope of the growth curve being 0.358. In contrast, the growth slope of the rats given the 1.25 micrograms dose of AXP was 0.621, whereas those given the CSF was 0.823. Although the trends of intake of food tended to follow the curves of the rats body weight, the difference between g/kg food intake of rats during ICV infusions of either dose of AXP was not significantly different from that of the CSF controls. Water intake also was unaffected by either dose of AXP. These results demonstrate that this tri-peptide derived from urine of patients afflicted with anorexia nervosa exerts a direct effect on the brain. Since the 0.25 micrograms dose of AXP infused acutely ICV caused a sustained hyperthermia, its mechanism of action is apparently a metabolic one; that is, the interruption in the gain in body weight of the rat is independent of the amount of food it ingests.

Animals↗