Luteinizing hormone-releasing hormone (LHRH) is a major regulator of reproduction in mammals. This decapeptide is processed from prohormone, pro-LHRH. At least four different enzymatic steps are involved in the conversion of pro-LHRH to LHRH. One of these processing enzymes may be carboxypeptidase E (CPE). Recently, a mouse line was identified with a point mutation in CPE. These CPEfat mice are obese, diabetic, and infertile. The overall objective of the proposal is to determine whether CPE is involved in processing the pro-LHRH in vivo and whether a defect in this enzyme is responsible for the infertility in the CPEfat mouse.
Aim I : There are four aspects to this first specific aim. First, processing of pro-LHRH will be examined in hypothalami from male and female wild type, heterozygous, and homozygous CPEfat mice. These data should identify which steps in pro-LHRH processing are deficient in the homozygotes. Second, LHRH gene expression and pro-LHRH biosynthesis will be compared among the groups of mice. Third, since pro-LHRH processing may involve three different enzymes besides CPE, expression of these enzymes will also be studied. Fourth, biological activity of the various pro-LHRH intermediates will be assessed in dispersed anterior pituitary cultures from these groups of mice. Finally, the hypothalamus, pituitary and gonads will be evaluated to determine whether the mutation in CPE affects additional responses from these organs.
Aim II : A transgenic approach will be used to rescue the CPEfat phenotype and restore processing of the pro-LHRH to LHRH. In this case, mice will be genetically targeted for CPE expression in hypothalamic LHRH neurons. These transgenic mice will be bred with heterozygous CPEfat mice to produce a mouse that is homozygous for the CPEfat mutation, but has CPE selectively expressed in LHRH neurons. The abilities of these mice to process pro-LHRH to LHRH and responses from the gonads, pituitary and hypothalamus will be evaluated as in Aim I.
Aim III : Mice homozygous for the CPEfat mutation that have selective expression of CPE in LHRH neurons will be evaluated for their abilities to reproduce. This series of experiments in the CPEfat mouse presents us with the unique opportunity to clearly link a deficiency in pro-LHRH processing with a defect in reproduction. The biochemical, physiological, and gene-targeting principles derived from this work should broaden our understanding of basic mechanisms that regulate reproduction in mammals and it may serve as a useful example for repairing genetically-based diseases or other defects in the future.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD036015-03
Application #
6182544
Study Section
Endocrinology Study Section (END)
Program Officer
De Paolo, Louis V
Project Start
1998-04-01
Project End
2003-03-31
Budget Start
2000-04-01
Budget End
2001-03-31
Support Year
3
Fiscal Year
2000
Total Cost
$209,129
Indirect Cost
Name
Duke University
Department
Psychiatry
Type
Schools of Medicine
DUNS #
071723621
City
Durham
State
NC
Country
United States
Zip Code
27705
Cheng, Y; Rodriguiz, R M; Murthy, S R K et al. (2015) Neurotrophic factor-?1 prevents stress-induced depression through enhancement of neurogenesis and is activated by rosiglitazone. Mol Psychiatry 20:744-54
Rodriguiz, Ramona M; Wilkins, John J; Creson, Thomas K et al. (2013) Emergence of anxiety-like behaviours in depressive-like Cpe(fat/fat) mice. Int J Neuropsychopharmacol 16:1623-34
Rodriguiz, Ramona M; Colvin, Jennifer S; Wetsel, William C (2011) Neurophenotyping genetically modified mice for social behavior. Methods Mol Biol 768:343-63
Srinivasan, Sudha; Bunch, Donna O; Feng, Yun et al. (2004) Deficits in reproduction and pro-gonadotropin-releasing hormone processing in male Cpefat mice. Endocrinology 145:2023-34
Cawley, Niamh X; Zhou, Jiechun; Hill, Joanna M et al. (2004) The carboxypeptidase E knockout mouse exhibits endocrinological and behavioral deficits. Endocrinology 145:5807-19
Longo, Maurizio; Brama, Marina; Marino, Maria et al. (2004) Interaction of estrogen receptor alpha with protein kinase C alpha and c-Src in osteoblasts during differentiation. Bone 34:100-11
Wetsel, William C; Srinivasan, Sudha (2002) Pro-GnRH processing. Prog Brain Res 141:221-41
Kreda, S M; Sumner, M; Fillo, S et al. (2001) alpha(1)-adrenergic receptors mediate LH-releasing hormone secretion through phospholipases C and A(2) in immortalized hypothalamic neurons. Endocrinology 142:4839-51
Xu, F; Gainetdinov, R R; Wetsel, W C et al. (2000) Mice lacking the norepinephrine transporter are supersensitive to psychostimulants. Nat Neurosci 3:465-71