Three phases of TRH-induced facilitation of exocytosis by single lactotrophs

Membrane capacitance measurements were used to study neuropeptide modulation of exocytosis by perforated patch clamped rat lactotrophs. We report that depolarizing voltage-clamp pulses evoke exocytosis that is steeply dependent on Ca²⁺ influx through voltage-gated Ca²⁺ channels. Furthermore, we find that the neuropeptide TRH (thyrotropin-releasing hormone) acts in three phases to promote exocytosis. First, TRH transiently (within ~0.5 min) triggers depolarization- and extracellular Ca²⁺- independent exocytosis. Second, within 3 min of application, TRH facilitates depolarization-evoked exocytosis while inhibiting voltage-gated Ca²⁺ current. Finally, after 8 min, TRH further enhances depolarization-evoked exocytosis by increasing high-voltage-activated (HVA) Ca²⁺ channel current. Activation of protein kinase C (PKC) with a phorbol ester also stimulates depolarization-evoked exocytosis by increasing Ca²⁺ current. Therefore, PKC can only account for the last effect of TRH. Thus, a single neuromodulator may employ several temporally distinct mechanisms to stimulate peptide secretion.