Culture of Enzymatically Dispersed Anterior Pituitary Cells: Functional Validation of a Method

Abstract

A method for preparing short—term enzymatically dispersed pituitary cell cultures is described. It has been used to assay hypothalamic releasing factors. The cells secrete TSH or LH and FSH in amounts directly related to the dose of TRF or LRF respectively. The minimal active doses of synthetic TRF and either synthetic or highly purified ovine LRF is ca. 10-10M. Apparent affinity constants for TRF or LRF receptors are ca. 10-9M. Enhanced secretion rates of TSH or LH of 8– to 20–fold in response to maximal levels of TRF or LRF are routinely observed. The sensitivity of the cultured cells to TRF and LRF is maintained for more than 2 weeks in culture although the magnitude of the secretory response and the intracellular hormone content declines with time in culture. Chronic administration of TRF or LRF depletes the intracellular hormone levels while simultaneously increasing the total TSH or LH content in the culture (tissue and fluid). As has been observed with other in vitro systems, high medium potassium, prostaglandin E2 and theophylline stimulate TSH secretion from the cultured cells. Furthermore, thyroid hormones which suppress the TSH secretion in response to TRF by the anterior pituitary in vivo and in other in vitro methods are similarly effective on these cells. The use of this technique has been applied to pituitary tissues of other species and types of cells, e.g., cells from G1 mouse thyrotropic tumors were shown to respond appropriately to TRF and thyroxine. The enzymatically dispersed cultured cells have been found to behave as would have been expected on the basis of experience with other in vitro methods while having significant advantages over those other procedures. (Endocrinology91: 562, 1972)