Secretory and radioligand binding studies on muscarinic receptors in bovine and feline chromaffin cells.

Abstract

SUMMARY1. Muscarinic agonists enhanced catecholamine release from perfused cat adrenal glands with the following relative order of potencies: methacholine > oxotremorine > McN-A-343 > pilocarpine > bethanechol > muscarine. Because a continuous online electrochemical detection system was used to monitor catecholamine release, this sequence could be obtained at concentrations much lower (1-10 /IbM) and during much shorter stimulation times (3-30 s) than in previous reports.2. All muscarinic agonists used secreted adrenaline preferentially over noradrenaline. Methacholine evoked a sustained, non-desensitizing response in the cat adrenal, which declined to basal levels of secretion immediately after Ca2+ removal; upon Ca2+ restoration secretion was restored to the previous plateau.3. In addition to evoking a direct secretory response, low concentrations of methacholine, pilocarpine, bethanechol or muscarine clearly potentiated cat adrenal secretory responses evoked by pulses of nicotine (2 4ttM 7. Because the ionophore A23187 enhanced K+-evoked secretion in both, bovine and cat adrenals, it seems that a similar cytosolic Ca2+ rise induced by muscarinic stimulation might constitute the underlying mechanism both to cause a secretory response per se as well as the potentiation of catecholamine release evoked by nicotinic or high K+ stimulation. However, it is unclear why the bovine behaves differently from the feline chromaffin cell as far as the muscarine-evoked effects are concerned. These differences might be explained by the fact that the cat adrenal chromaffin cell contains a single homogeneous population of muscarinic receptors of the M2 subtype, which seem to facilitate external Ca2+ entry through an associated ionophore. However, in the bovine adrenomedullary plasma membranes, two populations of muscarinic sites with different kinetic characteristics to those found in the cat might explain the different behaviour of muscarinic agonists.

Muscarinic receptor is coupled with a cation channel through a GTP‐binding protein in guinea‐pig chromaffin cells.

“…1965;Ballesta et al 1989), rat (Yoshizaki, 1973;Wakade & Wakade, 1983) and guinea-pig adrenal medulla (Role & Perlman, 1983), both type of receptors were found to be involved in the ACh-induced release of catecholamine. In fact, muscarinic activation of the cat adrenal medulla preferentially released adrenaline rather than noradrenaline whereas nicotinic stimulation of the same preparation secreted both hormones equally (Douglas & Poisner, 1965;Ballesta et al 1989). Despite such strong evidence for a physiological role of the muscarinic receptor in certain species of animals, the mechanisms underlying muscarinicinduced secretion still remain to be explored.…”

“…This ACh-induced secretion has been reported to be mediated through either the nicotinic receptor or the muscarinic receptor, or both, depending on the species of animal studied. Thus, the secretion of catecholamine from bovine chromaffin cells was entirely due to activation of the nicotinic receptor (Schneider, Herz & Rosenheck, 1977;Ballesta, Borges, Garcia & Hidalgo, 1989), whereas that from chicken adrenal medullary cells was caused by stimulation of the muscarinic receptor (Knight & Baker, 1986). In the cat (Douglas & Poisner.…”

“…These subtypes have been found to be distributed selectively over brain and peripheral organs, and are assumed to be coupled with specific functions. The muscarinic receptor in the cat adrenal medulla was reported to be of the M2 subtype from a study with pirenzepine displacement of labelled quinuclidyl benzilate methyl chloride (QNB) binding (Ballesta et al 1989). However, that was not the case in bovine chromaffin cells (Ballesta et al 1989).…”

“…The muscarinic receptor in the cat adrenal medulla was reported to be of the M2 subtype from a study with pirenzepine displacement of labelled quinuclidyl benzilate methyl chloride (QNB) binding (Ballesta et al 1989). However, that was not the case in bovine chromaffin cells (Ballesta et al 1989). The present experiment was undertaken to clarify muscarinic actions and pharmacological properties of the muscarinic receptor involved using isolated chromaffin cells of the guinea-pig.…”

Ca2+ Imaging in Perfused Adrenal Medullae

“…Stimulating electrodes are placed on the gland, and nerve fibers in the gland are transmurally stimulated. Catecholamines secreted from AM cells are measured by the f luorometric methods [2,3], the amperometric method [4] or high pressure liquid chromatography [5,6]. One advantage of this method would be that one can measure cellular activity from many AM cells.…”