WITHIN THE BODY, A PART OF THE BRAIN CALLED THE HYPOTHALAMUS RELEASES THE HORMONE, GONADOTROPIN RELEASING HORMONE (GNRH). THIS ACTS ON THE PITUITARY TO RELEASE LUTEINIZING HORMONE (LH), WHICH ACTS ON THE OVARIES TO RELEASE ESTRADIOL (E2). E2THEN FEEDS BACK IN A NEGATIVE MANNER AND INHIBITS THE RELEASE OF GNRH AND LH. IN MANY MAMMAL SPECIES, AS THEY REACHPUBERTY, THERE IS A DECREASE IN THE INHIBITION BY E2, LEADING TO AN INCREASE OF GNRH AND LH, WHICH EVENTUALLY LEADS TO PUBERTY ONSET. IT IS CURRENTLY UNKNOWN WHAT CAUSES THIS CHANGE IN E2-NEGATIVE FEEDBACK, AND THEREFORE WHAT CONTROLS THE TIMING OF PUBERTY ONSET. ADDING TO THE COMPLEXITY, E2CANNOT ACT DIRECTLY TOSUPPRESS GNRH, SINCE THE NEURONS THAT RELEASE GNRH LACK RECEPTORS FOR ESTRADIOL, ERA. THIS MEANS THAT THERE MUST BE OTHER NEURONS THAT HAVEERA AND WHICH ACTON GNRH NEURONS.DELAYEDPUBERTY ONSET IN DOMESTIC ANIMALS LEADS TO A DECREASE IN LIFETIME PRODUCTIVITY OF THE ANIMAL, THEREFORE DECREASING PROFITABILITY FOR PRODUCERS. IN ADDITION, DELAYED PUBERTY IN HUMANS HAS A NUMBER OF NEGATIVE EFFECTS ON HEALTH. DETERMINING WHAT MECHANISMS CONTROL THE TIMING OF PUBERTY ONSET MAY ALLOW FOR MORE OPTIONS TO ADDRESS THESE ISSUES.THE FIRST GOAL OF THIS PROJECT IS TO DETERMINE WHERE IN THE HYPOTHALAMUS ESTRADIOL IS ACTING. THE FIRST EXPERIMENT INVOLVES PLACINGE2IN SPECIFIC AREAS OF THE HYPOTHALAMUS, NAMELY EITHER THE PREOPTIC AREA (POA) OR THE ARCUATE NUCLEUS (ARC), OF EWESAFTER REMOVING THE OVARIES, AND THEREFORE ANY CIRCULATING ESTRADIOL. BLOOD SAMPLES WILL THEN BE TAKEN TO DETERMINE IF THE E2IMPLANTS REDUCE LH, INDICATING THAT E2IS ACTING IN EITHER THE POA OR ARC (OR BOTH), OR IF THERE IS NO RESPONSE. THE SECOND EXPERIMENT WILL INVOLVE PLACING ICI, A COMPOUND THAT BLOCKS THE RECEPTOR FOR E2, IN EITHER THE POA OR ARC OF EWES. THE OVARIES OF THE EWES WILL BE REMOVED, BUT E2WILL BE GIVENSO THAT THERE IS A CONSTANT AMOUNT OF ESTRADIOL IN CIRCULATION. BLOOD SAMPLES WILL BE TAKEN TO DETERMINE IF THE RESULTS SUPPORTTHE FIRST EXPERIMENT, I.E., IF E2IS ACTING IN THAT AREA, THE ICI COMPOUND SHOULD BLOCK IT FROM WORKING AND THE LH LEVELS WILL BE INCREASED.THE SECOND GOAL OF THE PROPOSAL IS TO TRY TO DETERMINE THE NEURONS IN THE HYPOTHALAMUSTHAT ACT ONGNRH NEURONS AND THATMAY BE RESPONSIBLE FOR THE CHANGE IN E2-NEGATIVE FEEDBACK THAT LEADS TO PUBERTY. WE HAVE IDENTIFIED A FEW POPULATIONS OF CELLS THAT WE KNOW ACT ON GNRH IN SOME WAY: ARC PROPIOMELANTOCORTIN (POMC) NEURONS, ARC AGOUTI-RELATED PEPTIDE (AGRP) NEURONS, AND POA NK3R-CONTAINING NEURONS. POMC AND AGRP NEURONS ARE KNOWN TO HAVE ERA RECEPTORS. APPROACHES WILL BE USED THAT WILL ENABLE US TO ASSESS CHANGES IN THE NUMBER OFPOMC, AGRP AND NK3R NEURONS WITH PUBERTAL DEVELOPMENT AS WELL AS WHETHER ERA EXPRESSION IN EACH OF THESE DIFFERENT SETS OF NEURONS CHANGES.IF ERA EXPRESSION ONE OR MORE OF THESE SETS OF NEURONS CHANGES, THEN IT MIGHT SUGGEST THAT THEY ARE INVOLVED IN THE CHANGE IN E2-NEGATIVE FEEDBACK THAT LEADS TO PUBERTY ONSET.