LACTOTROPHS SECRETING SMALL AMOUNTS OF PROLACTIN REVEAL GREAT RESPONSIVENESS TO THYROTROPIN-RELEASING-HORMONE - ANALYSIS BY THE SEQUENTIAL CELL IMMUNOBLOT ASSAY

The effects of TRH on PRL secretion from individual lactotrophs of female rats were investigated by using a sequential cell immunoblot assay. The same pituitary cells cultured on coverslips were first incubated to determine basal secretion of PRL and subsequently challenged by one of various concentrations of TRH. The PRL secreted from the single lactotrophs was absorbed on protein-blotting transfer membranes, immunostained, and quantified by microscopic image analysis. When no TRH was added to the medium used in the second incubation (controls), the amount of PRL secreted from individual lactotrophs was 93% of that secreted in the first incubation. Treatment with 3 x 10(-10)-10(-7) M TRH in the second incubation increased in a dose-dependent manner the proportion of lactotrophs whose PRL secretion was significantly greater than confidence limits for PRL secretion in the controls. However, the percentage of TRH-responsive lactotrophs remained less than 50% even at a maximally effective concentration of TRH. Proportions of the TRH-responsive lactotrophs were significantly greater in cells that secreted small amounts of PRL under basal conditions than in those that secreted large amounts. Furthermore, the small, but not the large, secretors showed a significant increase in mean absolute amounts of PRL secreted by 10(-7) M TRH, which represented no less than 45% of all PRL secreted from the lactotroph population by the TRH treatment. These results indicate that 1) there is a heterogeneity with respect to lactotroph responsiveness to TRH and that 2) a population of lactotrophs that secrete small amounts of PRL under basal conditions contains a much larger proportion of TRH-responsive lactotrophs than does a population of lactotrophs that secrete large amounts of PRL.