Ca(2+) and frequency dependence of exocytosis in isolated somata of magnocellular supraoptic neurones of the rat hypothalamus.

Abstract

In addition to action potential-evoked exocytotic release at neurohypophysial nerve terminals, the neurohormones arginine vasopressin (aVP) and oxytocin (OT) undergo Ca(2+)-dependent somatodendritic release within the supraoptic and paraventricular hypothalamic nuclei. However, the cellular and molecular mechanisms that underlie this release have not been elucidated. In the present study, the whole-cell patch-clamp technique was utilized in combination with high-time-resolved measurements of membrane capacitance (C(m)) and microfluorometric measurements of cytosolic free Ca(2+) concentration ([Ca(2+)](i)) to examine the Ca(2+) and stimulus dependence of exocytosis in the somata of magnocellular neurosecretory cells (MNCs) isolated from rat supraoptic nucleus (SON). Single depolarizing steps (> or =20 ms) that evoked high-voltage-activated (HVA) Ca(2+) currents (I(Ca)) and elevations in intracellular Ca(2+) concentration were accompanied by an increase in C(m) in a majority (40/47) of SON neurones. The C(m) responses were composed of an initial Ca(2+)-independent, transient component and a subsequent, sustained phase of increased C(m) (termed DeltaC(m)) mediated by an influx of Ca(2+), and increased with corresponding prolongation of depolarizing step durations (20-200 ms). From this relationship we estimated the rate of vesicular release to be 1533 vesicles s(-1). Delivery of neurone-derived action potential waveforms (APWs) as stimulus templates elicited I(Ca) and also induced a DeltaC(m), provided APWs were applied in trains of greater than 13 Hz. A train of APWs modelled after the bursting pattern recorded from an OT-containing neurone during the milk ejection reflex was effective in supporting an exocytotic DeltaC(m) in isolated MNCs, indicating that the somata of SON neurones respond to physiological patterns of neuronal activity with Ca(2+)-dependent exocytotic activity.