In situ hybridization detection of marked differences in pre-proopiomelanocortin messenger ribonucleic acid content of individual corticotropes and melanotropes.
Recently, heterogeneity of POMC mRNA content between intermediate lobe melanotropes of the rat pituitary gland was demonstrated by in situ hybridization of tissue sections. In the present study the heterogeneity of POMC mRNA content in dispersed rat pituitary cells has been investigated. Acutely dispersed cells from adult male rat anterior or neurointermediate lobe tissues were adhered to poly-L-lysine-coated coverslips. The cells were fixed and then hybridized with 35S-labeled POMC or 1B15 (cyclophilin) cRNA. Parallel studies measuring constitutively expressed cellular 1B15 mRNA content were undertaken to ensure that the apparent single cell differences in POMC mRNA were not inherent to the in situ hybridization procedure. When classified by image analysis, extensive differences in silver grain densities were seen over POMC mRNA-containing cells from both lobes. To determine if mRNA in polysomal configurations was less accessable for hybridization with probes than naked mRNA, cells were preincubated with pactamycin, a potent inhibitor of ribosomal initiation of protein synthesis. Pactamycin had no effect on these results. Thus, there appears to be large differences in POMC mRNA content between individual pituitary cells expressing the same gene product.
Hatfield
J M
JM
Department of Biochemistry, Oregon Health Sciences University School of Medicine, Portland 97201.
Daikh
D I
DI
Adelman
J P
JP
Douglass
J
J
Bond
C T
CT
Allen
R G
RG
D000324:Adrenocorticotropic Hormone / Q000378:metabolism*; D000818:Animals; D008297:Male; D009074:Melanocyte-Stimulating Hormones / Q000378:metabolism*; D009693:Nucleic Acid Hybridization; D010142:Pactamycin / Q000494:pharmacology; D010902:Pituitary Gland / Q000032:analysis* / Q000166:cytology / Q000187:drug effects; D010903:Pituitary Gland, Anterior / Q000032:analysis / Q000166:cytology; D011333:Pro-Opiomelanocortin / Q000235:genetics*; D015347:RNA Probes; D012333:RNA, Messenger / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains
D015347:RNA Probes; D012333:RNA, Messenger; D010142:Pactamycin; D011333:Pro-Opiomelanocortin; D000324:Adrenocorticotropic Hormone; D009074:Melanocyte-Stimulating Hormones