Acetylcholine, released from cholinergic nerve terminals, innervated to adrenal chromaffin cells, evokes catecholamine release from the chromaffin cells. In the previous studies neostigmine, an acetylcholinesterase inhibitor, nicotine and oxotremorine, a muscarinic receptor agonist, hardly induce catecholamine release from adrenal medulla of 21-day-old rats. Not only adrenaline but also noradrenaline is released from the chromaffin cells by insulin-induced hypoglycemia in fasted 21-day-old rats, whereas preferential adrenaline release occurs in fasted 8-week-old rats. The purpose of the present study was to characterize the catecholamine output induced by hypoglycemia in 21-day-old rats. Hexamethonium, a nicotinic receptor antagonist, blocked the adrenaline and noradrenaline release from the chromaffin cells almost completely as judged from measuring the catecholamine content and observing the morphological changes of the chromaffin cells. Nicotine or oxotremorine, injected into the fasted animals, induced catecholamine release. In the multiple steps of the chromaffin granule exocytotic pathway one or several steps are probably inactive in infant rat and these steps become to be fully active by starvation. The nicotinic receptors of the chromaffin cells in infant rats mainly contribute the secretion induced by hypoglycemia. It has been postulated that the nicotinic receptors are primarily concentrated in the synaptic zones of the chromaffin cell membrane and are involved in the physiological stimulation, whereas the extrasynaptic regions contain a mixture of the nicotinic and muscarinic receptors, these are activated by injected secretogogues.