Calcium signalling in single growth hormone-releasing factor-responsive pituitary cells

The release of pituitary GH appears to be critically dependent on alterations in the free intracellular Ca²⁺ concentration ([Ca²⁺]_i). However, little is known about the nature of Ca²⁺ signalling within normal pituitary cells We, therefore, examined [Ca²⁺]_i patterns in individual cultured pituicytes of adult male rats under basal conditions and in response to GH regulatory agents, using the calcium-sensitive dye fura-2 together with digital imaging microscopy. Perfusion of cultured anterior pituitary cells with GH-releasing factor (GHRF) resulted in a marked increase in [Ca²⁺]_i in specific pituitary cells. These cells did not respond to other hypothalamic secretagogues (GnRH, TRH, or CRF), and there was no evidence of desensitization on repetitive administration of GHRF. Somatotrophs (n = 134) exhibited spontaneous oscillations of [Ca²⁺]_i in the basal state, with considerable heterogeneity of oscillatory patterns among cells. After application of a near maximal stimulatory dose of GHRF (1 nM), there was a striking 2.2-fold increase in the amplitude of [Ca²⁺]_i oscillations and only a modest increase in their frequency. Forskolin (1 μM) augmented somatotroph [Ca²⁺]_i in patterns similar to those of GHRF. Somatostatin (10 nM) abolished the [Ca²⁺]_i response to GHRF (n = 26); this reflected a marked reduction in the amplitude of [Ca²⁺]_i oscillations and a slight reduction in their frequency. Ca²⁺-free medium or the Ca²⁺ channel antagonist nimodipine (0.1-1 μM) suppressed the Ca²⁺ stimulatory effect of GHRF. Conversely, the Ca²⁺ channel agonist BAY K8644 (1 μM) strikingly augmented the GHRF-induced rise in [Ca²⁺]_i, with a major stimulatory effect on the amplitude of [Ca²⁺]_i oscillations and no observed effect on their frequency. In summary, GHRF and other hypothalamic secretagogues increase [Ca²⁺]_i in pituitary cells in a highly specific manner, consistent with the known specificity of their effects on hormone release. Somatotrophs exhibit spontaneous rhythmic oscillation of [Ca²⁺]_i in the basal state. Known regulators of GH release markedly alter the [Ca²⁺]_i oscillatory pattern in characteristic manners, exerting predominant effects on the amplitude of [Ca²⁺]_i pulses and lesser effects on their frequency. These striking effects of GH regulatory agents on pituitary Ca²⁺ signalling are consistent with the concept that modulation of [Ca²⁺]_i is a critical mediator of somatotroph function.