OBJECTIVE: Proopiomelanocortin (POMC) gene is involved in various neuroendocrine reproductive activities and its principal areas of expression are hypothalamus and pituitary gland. However, as it is known as low copy gene, detection of its expression by in situ hybridization is challenging. This study was to examine the degree of basal expression of POMC mRNA and anatomic localization of its expression in rat hypothalamus and pituitary gland as a preliminary study for related reproductive endocrine researches. METHODS: Hypothalamus and pituitary gland of ovariectomized female Spraque-Dawley rat (weight: 250-300 g) was obtained by decapitation with preservation of POMC mRNA by cardiac perfusion of 4% paraformaldehyde. Then, coronal sections of hypothalamus and horizontal sections of pituitary gland were made after paraffin embedding. For in situ hybridization, 35S-labeled antisense RNA probe was used and after in situ hybridization reaction and serial washing procedure, detection of its signal was made by emulsion autoradiography of slide and dark field microscopic examination was done for photographic documentation. Anatomic localization of various nucleus of hypothalamus was done by method proposed by Paxinos and Watson1. RESULTS: Strong expression signal of POMC mRNA was detected in the intermediate lobe of pituitary gland. Less strong, but evenly distributed signals were also detected in the anterior lobe of pituitary gland. In hypothalamic sections, entire area of arcuate nucleus from rostral to caudal direction (between bregma -2.80 mm to bregma -4.30 mm area) showed clear and strong signal of POMC mRNA expression. CONCLUSION: Though POMC gene is known as low copy gene, its basal mRNA expression in pituitary gland and hypothalamus in ovariectomized rat were detectable by in situ hybridization technique. As subpopulation of arcuate nucleus are under complex differential regulation, this in situ hybridization model would be helpful for understanding the effects of various regulatory factors on expression of POMC gene in the specific anatomic area of hypothalamus and pituitary gland.
Animals ; Arcuate Nucleus ; Autoradiography ; Decapitation ; Female ; Humans ; Hypothalamus* ; In Situ Hybridization* ; Paraffin Embedding ; Perfusion ; Pituitary Gland* ; Pituitary Gland, Anterior ; Pituitary Gland, Intermediate ; Pro-Opiomelanocortin* ; Rats* ; RNA, Antisense ; RNA, Messenger