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Functional Characterization of Genetically Labeled Gonadotropes

Institute for Neural Signal Transduction (S.W., J.R.S., D.N., C.D., W.H., U.B.), Center for Molecular Neurobiology, D-20253 Hamburg, Germany; and Institute for Vegetative Physiology and Pathophysiology (C.K.B.), University Hospital Hamburg-Eppendorf, D-20246 Hamburg, Germany

Address all correspondence and requests for reprints to: Dr. Ulrich Boehm, Institute for Neural Signal Transduction, Center for Molecular Neurobiology, Falkenried 94, D-20253 Hamburg, Germany. E-mail: ulrich.boehm{at}zmnh.uni-hamburg.de.

Gonadotropes are crucial in the control of reproduction but difficult to isolate for functional analysis due to their scattered distribution in the anterior pituitary gland. We devised a binary genetic approach, and describe a new mouse model that allows visualization and manipulation of gonadotrope cells. Using gene targeting in embryonic stem cells, we generated mice in which Cre recombinase is coexpressed with the GnRH receptor, which is expressed in gonadotrope cells. We show that we can direct Cre-mediated recombination of a yellow fluorescent protein reporter allele specifically in gonadotropes within the anterior pituitary of these knock-in mice. More than 99% of gonadotropin-containing cells were labeled by yellow fluorescent protein fluorescence and readily identifiable in dissociated pituitary cell culture, allowing potentially unbiased sampling from the gonadotrope population. Using electrophysiology, calcium imaging, and the study of secretion on the single-cell level, the functional properties of gonadotropes isolated from male mice were analyzed. Our studies demonstrate a significant heterogeneity in the resting properties of gonadotropes and their responses to GnRH. About 50% of gonadotropes do not exhibit secretion of LH or FSH. Application of GnRH induced a broad range of both electrophysiological responses and increases in the intracellular calcium concentration. Our mouse model will also be able to direct expression of other Cre recombination-dependent reporter genes to gonadotropes and, therefore, represents a versatile new tool in the understanding of gonadotrope biology.

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