Ob receptor in rabbit ovary and leptin in vitro regulation of corpora lutea.
We studied leptin involvement in rabbit corpora lutea (CL) activity, and its post-transcriptional signalling pathway. The expression of leptin receptor (Ob-R) in rabbit ovary at day 9 of pseudopregnancy was evaluated by immunohistochemistry and Western blot analysis. The specificity of the Ob-R receptor antibodies was characterised by immunoprecipitation and competition with blocking peptide. Day 9 CL were incubated in vitro with leptin alone or with inhibitors of PLC (phospholipase C), PLD (phospholipase D), AC (adenylate cyclase), JAK (janus kinase), MAPK (mitogen-activated protein kinase) and both cAMP- and cGMP-specific PDE (phosphodiesterase). Prostaglandin F2alpha(PGF2alpha), PGE2 and progesterone levels were measured in the culture medium, while NOS (nitric oxide synthase) and cAMP- and cGMP- specific PDE activities were measured in CL tissue. Positive staining for Ob-R was found within the cytoplasm of large luteal cells of CL as well as in granulosa cells of follicles and oocytes. Immunoblots detected a band of about 99 kDa size in Ob-R immunoprecipitates from CL homogenates. This band was not detectable after pre-incubation of the primary antibody with the immunising leptin peptide. Leptin increased PGF2alphaand cAMP-specific PDE, decreased basal progesterone and did not affect PGE2 and NOS levels. Leptin used the JAK pathway in increasing PGF2alpha, and MAPK and cAMP-specific PDE in decreasing progesterone. This study supports a permissive luteolytic role for leptin in rabbit CL.
Zerani
Massimo
M
Dipartimento di Biologia molecolare, cellulare e animale, Università di Camerino, via F. Camerini 1, 62032 Camerino, Italy. massimo.zerani@unicam.it
Boiti
Cristiano
C
Zampini
Danilo
D
Brecchia
Gabriele
G
Dall'Aglio
Cecilia
C
Ceccarelli
Piero
P
Gobbetti
Anna
A
D000818:Animals; D015153:Blotting, Western / Q000379:methods; D003338:Corpus Luteum / Q000187:drug effects*; D015237:Dinoprost / Q000032:analysis; D005260:Female; D007150:Immunohistochemistry / Q000379:methods; D020738:Leptin / Q000494:pharmacology*; D019001:Nitric Oxide Synthase / Q000032:analysis; D010053:Ovary / Q000737:chemistry*; D010727:Phosphoric Diester Hydrolases / Q000032:analysis; D011374:Progesterone / Q000032:analysis; D011555:Pseudopregnancy; D011817:Rabbits; D011956:Receptors, Cell Surface / Q000032:analysis*; D054411:Receptors, Leptin; D046509:Tissue Culture Techniques
D020738:Leptin; D011956:Receptors, Cell Surface; D054411:Receptors, Leptin; D011374:Progesterone; D015237:Dinoprost; D019001:Nitric Oxide Synthase; D010727:Phosphoric Diester Hydrolases