Magnocellular neurons of the hypothalamic supraoptic nucleus have been shown to express the immediate-early gene c-fos in a number of experimental and physiological circumstances. In each case the induction of the immediate-early gene followed the increase in the spike activity of the cells. Since an increase in the intracellular concentration of calcium following influx through voltage-sensitive calcium channels is a known stimulus for c-fos expression and since the action potentials of these neurons have a large calcium component, we hypothesized that c-fos induction in these neurons could be attributed to calcium influx during spike activity. In the present experiments we use extracellular recording and immunocytochemistry for Fos, the protein product of c-fos, to demonstrate the activation of the cells following intracerebroventricular administration of the muscarinic agonist, carbachol. Fos expression following carbachol injection was then compared with that induced by a similar number of antidromically evoked action potentials. Antidromic activation, unlike the activation induced by carbachol, did not lead to the induction of Fos. We conclude that Fos induction in these neurons requires receptor activation rather than spike activity.