OPIOID MODULATION OF THE RELEASE OF LUTEINIZING HORMONE (PITUITARY, CELL CULTURE, DYNORPHIN, MET-ENKEPHALIN)
Abstract
A cell suspension culture of bovine anterior pituitary was validated for studying the release of luteinizing hormone (LH). Dissociation of the pituitary cells with higher concentrations of trypsin yielded more individual cells, but optimal LH secretion resulted from the use of concentrations of trypsin equivalent to 8-16 (mu)g/mg tissue for dissociation. Pituitary cells either freshly dissociated or preincubated for 18 h could be used to study LH secretory responses to gonadotropin-releasing hormone (GnRH) in subsequent 2 h experimental culture. After 18 h preincubation, experimental cultures lasting between 6 and 24 h could be used to study an inhibitory effect of a compound that required a longer exposure. Following cell dissociation, physiological concentrations of estrogen tended to augment total cellular LH and basal LH release after 18-h treatment but not after 2-h treatment. Progesterone had little effect by itself on pituitary LH in dispersed cells. Naloxone (1 (mu)M), an opioid antagonist, increased (P < .01) basal release of LH by 57% after 2 h culture but not after 24 h culture. Naloxone did not augment the amount of LH released in response to 10 nM GnRH. Methionine-enkephalin (Met-EK, 1 nM to 1 (mu)M) suppressed (P < .05) basal release of LH during both 2-h and 24-h cultures. Met-EK (1 and 100 nM) antagonized (P < .05) the stimulatory effect of naloxone suggesting an action specifically mediated by opioid receptors. Met-EK also attenuated (P < .05) the short-term stimulatory effect of GnRH. Different forms of immunoreactive dynorphin-A (ir-DYN-A) were present in the anterior lobe (AL) and neurointermediate lobe (NIL) of both bovine and ovine pituitaries. The AL contained a large molecular weight form of ir-DYN-A while the NIL contained primarily smaller forms of ir-DYN-A. Five-day treatment with steroids in vivo depressed AL concentration of ir-DYN-A while 2-h treatment of dispersed AL cells in vitro augmented the release of ir-DYN-A without altering pituitary LH. Eighteen-h preincubation restored pituitary contents of both LH and ir-DYN-A. Upon in vitro treatment with GnRH, dispersed AL cells increased their release of LH but not ir-DYN-A suggesting a dissociation between LH producing cells and ir-DYN-A-producing cells in the AL.
Degree
Ph.D.
Subject Area
Livestock
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