Targeting the Gonadotropin Releasing Hormone Neurons with Green Fluorescent Protein and Rodhamine
GnRH neurons form the final common pathway for central control of reproduction, with regulation achieved by changing the pattern of GnRH pulses. These neurons are diffusely scattered from the diagonal band of Broca throughout the medial basal hypothalamus thus making isolation of GnRH neurons nearly impossible. As a result, the molecular mechanisms underlying the physiological network of these neurons remain largely hypothetical. While previous studies have concluded that there are ~ 800 GnRH neurons located in the medial basal hypothalamus, recent studies suggest that the actual number may be much greater. To asses the number of GnRH neurons we used transgenic mice in which the green fluorescent protein reporter is genetically targeted for the expression of GnRH neurons allowing an 84-94% identification. Inmunohistochemistry was used in intact and ovariectomized to double stain the GnRH neurons with Rodhamine to allow for a more accurate count of GnRH neurons. The signal derived from the GnRH - GFP neurons was relatively strong, permitting us to delineate precisely the boundaries of the GnRH expressing cells. In 12 brain slices of the pre optic area of GFP-GnRH mice we counted approximately 1,000 - 1,200 GFP positive GnRH neurons. Furthermore, our current research suggests that the number of neurons that express GnRH, at detectable levels, changes when the animal's steroidal environment is altered. Isolating a pure population of GnRH neurons will allow us to perform molecular analysis to asses the effects of the hormonal environment, circadian timing, nutrition and stress on GnRH neurons. Before these ambitious goals can be tackled, however, we need to more completely understand the cells that constitute the population of GnRH neurons.
School:
University of Illinois at Urbana-Champaign
Department:
Human Biology
Research Advisor:
Shelley Tischkau
Department of Research Advisor:
Veterinary Biosciences
Year of Publication:
2005