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Galanin exerts an inhibitory effect on locus coeruleus (LC) neurons via a postsynaptic, as yet unidentified galanin receptor. Using an in vitro intracellular recording technique the effect of two galanin receptor agonists on LC neurons was investigated. Bath application of [Sar(1), D-Ala(12)]gal(1-16)-NH(2) (AR-M961), an agonist both at galanin R1 and R2 (GALR1, GALR2) receptors, evoked a reversible membrane hyperpolarization and inhibition of spike discharge in all LC neurons tested (n=42). The action of AR-M961 was blocked by tetraethylammonium chloride. Hyperpolarizing responses induced by AR-M961 were retained in the presence of tetrodotoxin and high Mg(2+)/low Ca(2+) media. The selective GALR2 agonist Gal(2-11)-NH(2) (AR-M1896) only caused inhibition of spike discharge and a slight hyperpolarization in 26 of 34 LC neurons tested, and was on a molar basis much weaker than AR-M961. These results suggest that it mainly is the GALR1 receptor that mediates hyperpolarization of LC neurons.

Cytoplasmic calcium concentration was measured in neurosecretory nerve terminals (neurosecretosomes) isolated from rat neurohypophyses by fura-2 fluorescence measurements and digital video microscopy. Hormone release and cytoplasmic calcium concentration were measured during depolarizations induced by elevated extracellular potassium concentration. During prolonged depolarizations with 55 mM [K+]o, the cytoplasmic calcium concentration remained elevated as long as depolarization persisted, while secretion inactivated after the initial sharp rise. The amplitude and duration of the increase in [Ca2+]i was dependent on the degree of depolarization such that upon low levels of depolarizations (12.5 mM or 25 mM [K+]o), the calcium responses were smaller and relatively transient, and with higher levels of depolarization (55 mM [K+]o) the responses were sustained and were higher in amplitude. Responses to low levels of depolarization were less sensitive to the dihydropyridine calcium channel blocker, nimodipine, while the increase in [Ca2+]i induced by 55 mM [K+]o became transient, and was significantly smaller. These observations suggest that these peptidergic nerve terminals possess at least two different types of voltage-gated calcium channels. Removal of extracellular sodium resulted in a significant increase in [Ca2+]i and secretion in the absence of depolarizing stimulus, suggesting that sodium-calcium exchange mechanism is operative in these nerve terminals. Although the [Ca2+]i increase was of similar magnitude to the depolarization-induced changes, the resultant secretion was 10-fold lower, but the rate of inactivation of secretion, however, was comparable.