Electrophysiological measurements of voltage-dependent Na+ influx using patch-clamp methodology were combined with optical monitoring of the free intracellular Na+ concentration in isolated rat neurohypophysial nerve endings to determine the relationship between Na+ influx generated by repetitive stimulation and change in [Na+]i. Application of step depolarizations under voltage-clamp-evoked tetrodotoxin-sensitive inward currents that were dependent upon extracellular Na+ and that exhibited rapid activation and inactivation properties. These characteristics substantiated the evoked current as a voltage-dependent Na+ current. Application of stimulus trains consisting of step depolarizations that mimick in frequency and duration those of action potentials were found to result in increases in [Na+]i. The induced change in [Na+]i was found to be related to the frequency and period of stimulation. Changes in [Na+]i were greatest at frequencies of 40 Hz and gave maximal changes with 30 s of continuous stimulation of approximately 2.4 mM. Sodium influx expressed as a molar quantity resulted in a nearly directly proportional increase in [Na+]i during the initial period of stimulation at low Na+ loads. When expressed as a charge density (pC/microm2) Na+ influx was found to increase with smaller diameter nerve endings as did the rate of change in [Na+]i in response to applied repetitive step depolarizations. Repetitive step depolarizations which simulate impulse activity that invade neuroendocrine nerve endings in vivo in response to physiological demand for hormone secretion resulted in an increased [Na+]i. It is postulated that this increased [Na+]i may provide a modulatory influence on the secretory response indirectly via alteration of intracellular calcium regulation or, perhaps, via a direct action on the secretory mechanism.