Effect of tetrodotoxin on adrenaline secretion in the perfused rat adrenal medulla

Abstract

EXTRACELLULAR Ca ions are required for the secretion of many hormones1. In the adrenal medulla catecholamine secretion evoked by acetylcholine (ACh) is completely blocked by omitting Ca from the medium2,3. The mechanism for Ca entry has been studied in a phaeochromocytoma cell line (PC12) which originated from a rat adrenal medullary tumour4,5. Two alternative ways of Ca entry were considered, voltage-dependent Ca channels and ACh channels. By analysing dopamine secretion4 and radioactive Ca influx5 in various ionic environments it was concluded that more than 80% of the Ca ions enter through the voltage-dependent Ca channels during stimulation by cholinergic agonists. Action potentials mediated by Na6,7 and Ca7 have been demonstrated in primary cultures of rat adrenal chromaffin cells. Furthermore, extracellular recording techniques can detect spontaneous action potentials in these cells. The frequency of these action potentials is modified in a dose-dependent manner by ACh (3×10−7M to 10−4M)7, and the same range of ACh concentrations stimulates adrenaline secretion in the perfused rat adrenal medulla3,8. Thus modulation of action potential frequency by ACh may be a mechanism for controlling the amount of Ca entry. The potential change during a Na action potential may activate voltage-dependent Ca channels, thereby increasing Ca influx. Also, spontaneous action potentials are blocked by 6 µM tetrodotoxin (TTX)7, a specific inhibitor of Na channels9. Therefore, if the Na action potential is involved in secretion, TTX should reduce Ca entry which in turn should decrease catecholamine secretion. We tested this hypothesis in the perfused rat adrenal medulla and report here that TTX decreases a portion of the adrenaline secretion evoked by either high KCl or ACh. Thus, Na action potentials, by facilitating the activation of Ca channels, may be involved in the regulation of adrenaline secretion from adrenal chromaffin cells.