PubMed Health⌕ Search

Biomedical subjects

L W Role

Publications and source records attributed to L W Role.

46 records · Page 3Linked to original sources
PubMed1985

On the mechanism of acetylcholine receptor accumulation at newly formed synapses on chick myotubes.

We have examined the specificity and the mechanism of acetylcholine receptor (AChR) accumulation at embryonic chick nerve-muscle contacts that form in culture. Spinal cord motoneurons were identified in vitro after labeling them in vivo with Lucifer Yellow-wheat germ agglutinin conjugates. All of their processes induced receptor clusters on contacted myotubes; after 24 to 48 hr of co-culture, the incidence of neurite-associated receptor patches (NARPs) was approximately 1.2/100 microns of contact. In contrast, NARPs were rarely associated with spinal cord interneurons (approximately 0.1/100 microns of contact). Neurons dissociated from ciliary ganglia induce NARPS to the same extent as motoneurons. The relative contribution to NARPs of AChRs present in the membrane prior to plating ciliary ganglion neurons and of "new" AChRs inserted 8, 11, or 17 hr after addition of neurons was assessed with two fluorescent receptor probes. Rhodamine-conjugated alpha-bungarotoxin was used to label either old or new receptors; a monoclonal, anti-receptor antibody visualized with fluorescein-second antibody was used to label all (new and old) receptors. Analysis of digitized fluorescence images showed that NARPs contained both new and old receptors but that within the first 24 hr of co-culture the majority (60 to 80%) were new. We estimate that cholinergic neurites increase the rate of receptor insertion 4- to 5-fold during the first 8 hr of NARP formation. The contribution of new receptors to NARPs declines with time. After 3 days of co-culture, receptors inserted over an 8-hr interval comprised only 20% of the total NARP complement.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Substance P modulation of acetylcholine-induced currents in embryonic chicken sympathetic and ciliary ganglion neurons.

Substance P has been identified by combined immunohistochemical and radioimmunological techniques to be present in preganglionic cholinergic and sensory nerve fibers of amphibian, mammalian, and avian autonomic ganglia. The peptide has been shown to depolarize sympathetic neurons of frog and guinea pig and to decrease the cholinergic activation of Na+ influx and catecholamine release from chromaffin cells. The aim of this study was to examine the interaction of acetylcholine and substance P on autonomic neurons. This report demonstrates a direct effect of substance P on acetylcholine-induced inward currents in both sympathetic and parasympathetic neurons clamped near resting membrane potential. Under these conditions, substance P dramatically enhances the rate of decay of the inward current in the continued presence of agonist without substantially affecting peak inward current. This effect is consistent with an enhancement of acetylcholine-receptor desensitization. Since substance P-containing cell bodies have been demonstrated in the avian (preganglionic) column of Terni as well as in fibers from the nucleus of Edinger-Westphal, the observed peptide inhibition of cholinergic activation of the neurons may function physiologically to modulate synaptic function in autonomic ganglia.

Acetylcholine↗
PubMed1983

Both nicotinic and muscarinic receptors mediate catecholamine secretion by isolated guinea-pig chromaffin cells.

We have studied the roles of nicotinic and muscarinic receptors in the acetylcholine-evoked secretion of catecholamine from guinea-pig chromaffin cells. Isolated guinea-pig chromaffin cells secrete catecholamine in response to acetylcholine, nicotine, and a variety of muscarinic agonists. Optimal concentrations of acetylcholine (50-200 microM) induce the release of 10-25% of the catecholamine content of the cells in 10 min. Maximal secretion evoked by nicotine or by muscarinic agonists is 5-12% of the catecholamine content of the cells. Secretion evoked by optimal concentrations of nicotine (50 microM) and muscarine (200 microM) are additive, and together these agonists cause catecholamine release equivalent to that produced by optimal concentrations of acetylcholine. Atropine causes a biphasic inhibition of acetylcholine-induced catecholamine secretion; low concentrations of atropine (0.02-0.01 microM) inhibit by 35-45% the catecholamine secretion evoked by 100 microM acetylcholine. Increasing the atropine concentration from 0.1 to 5 microM causes no further decrease in acetylcholine-evoked release, but at concentrations above 5 microM, a second distinct phase of inhibition appears. At 100 microM, atropine reduces acetylcholine-evoked secretion by 85%. At 0.1 microM, atropine significantly inhibits secretion induced by muscarinic, but not nicotinic, agonists. Tubocurarine (50 microM) does not block muscarinic stimulation of release, but inhibits acetylcholine- and nicotine-evoked release by 70 and 80%, respectively. Our experiments indicate that nicotinic and muscarinic stimulation represent distinct mechanisms for the activation of catecholamine release from guinea-pig chromaffin cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Catecholamine uptake into isolated adrenal chromaffin cells: inhibition of uptake by acetylcholine.

We have investigated the process of catecholamine uptake in guinea-pig chromaffin cells. Isolated guinea-pig chromaffin cells accumulate [3H]norepinephrine and [3H]epinephrine by a saturable transport system. Catecholamine uptake is dependent upon temperature, energy, and extracellular Na+. The apparent KmS for norepinephrine and epinephrine transport are approximately 1 and 3.5 microM, respectively; the transport maximum (Vmax) for both compounds is about 100 pmol/min/mg protein. The uptake of norepinephrine into chromaffin cells is inhibited by imipramine (Ki = 50 nM) and by desmethylimipramine (IC50 = 20 nM). In both its substrate specificity and its sensitivity to pharmacological inhibition, the catecholamine uptake system in chromaffin cells is similar to the catecholamine transport system previously described in sympathetic neurons. Decreasing external Na+ from 130 to 19 mM increases the apparent Km for norepinephrine to 2.8 microM. Decreasing external norepinephrine increases the Na+ concentration required for half-maximal transport. Agents that depolarize chromaffin cells, such as acetylcholine and veratridine, significantly inhibit [3H]norepinephrine uptake. This decrease in uptake is due to an increase in the apparent Km for norepinephrine. The inhibition of [3H]norepinephrine uptake by depolarizing agents cannot be accounted for by the preferential release of newly-accumulated [3H]norepinephrine, or by the competitive inhibition of [3H]norepinephrine uptake by secreted catecholamines. The inhibition of catecholamine uptake by depolarizing agents suggests that the transport system may be regulated by the membrane potential. Norepinephrine and epinephrine that are spontaneously released from the adrenal medulla may be recaptured in vivo. The inhibition of transport by acetylcholine may prevent the re-uptake of catecholamine released during the physiological stimulation of secretion.

Acetylcholine↗
PubMed1980

Purification of adrenal medullary chromaffin cells by density gradient centrifugation.

A method has been developed for the isolation of chromaffin cells from guinea pig adrenal glands. Crude suspensions of adrenal cells are prepared by the digestion of adrenal with collagenase. Chromaffin cells are then purified from these crude suspensions by isopycnic centrifugation through a 5-25% (w/v) gradient of metrizamide. More than 90% of the cells in the preparation are viable chromaffin cells, as judged by trypan blue exclusion and fluorescence histochemistry. A more convenient method for monitoring the purity of chromaffin cell preparations, using neutral red staining, is also described. Chromaffin cells are the primary neutral red staining cell in the adrenal gland, and more than 90% of the purified chromaffin cells stain with this dye. Purified chromaffin cells contain 400 +/- 50 nmol of epinephrine/mg protein (133 +/- 16 nmol/10(6) cells), and secrete epinephrine in response to acetylcholine. At a concentration of 100 microM, acetylcholine causes a 10- to 20-fold increase in the secretion of epinephrine from the cells. The method described in this paper is a useful procedure for the preparation of pure, functional chromaffin cells.

Adrenal Medulla↗
PubMed1980

Mechanisms of ionophore-induced catecholamine secretion.

A number of carboxylic ionophores stimulate the secretion of norepinephrine from cell suspensions prepared from a transplantable rat pheochromocytoma. The divalent-cation ionophore ionomycin stimulates catecholamine secretion by a mechanism that is dependent upon the presence of extracellular Ca++. It is likely that ionomycin-induced catecholamine secretion results from the ionophore-mediated entry of Ca++ into the cells. The monovalent-cation ionophore monensin stimulates catecholamine secretion by a mechanism that is independent of extracellular Ca++, but is markedly dependent upon extracellular Na+. Monensin probably transports Na+ into the pheochromocytoma cells and increases the intracellular concentration of Na+ in these cells. This rise in intracellular Na+ may cause the release of Ca++ from some intracellular store. Lasalocid stimulates catecholamine secretion by a mechanism that is independent of extracellular Ca++ and is only slightly dependent upon extracellular Na+. The action of lasalocid, in contrast to the actions of ionomycin and monensin, is potentiated by decreased pH. It is likely that lasalocid enters the cells in its uncharged, protonated form. Once inside the cells, lasalocid may promote the release of intracellular Ca++. Alternatively, lasalocid and monensin may stimulate catecholamine secretion by the process which is independent of Ca++. These experiments show that ionophores can stimulate catecholamine secretion by at least three distinct ionic mechanisms.

Adrenal Gland Neoplasms↗
PubMedDate not supplied

Acetylcholine release from growth cones detected with patches of acetylcholine receptor-rich membranes.

Synaptic potentials can be evoked at nerve-muscle junctions in vitro within minutes after an exploring growth cone contacts a receptive myotube. Functional transmission is also evident in vivo on the time scale of minutes after motor axons enter adjacent myotomes. The ability to release acetylcholine (ACh) may be induced in motor nerve terminals after they contact receptive target cells. Alternately, growth cones may be capable of releasing ACh before contact. To examine the development of ACh release we have used isolated patches of acetylcholine receptor(AChR)-rich membrane as sensitive detectors of ACh. We report here that the growth cones of embryonic chick ciliary ganglion neurones can release ACh, even when the cells are grown in the absence of target myotubes.

Acetylcholine↗