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 which 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 YFP reporter allele specifically in gonadotropes within the anterior pituitary of these knock-in mice. More than 99% of gonadotropin containing cells were labeled by YFP 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.