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Updated: Oct 23, 2026
Immunohistochemical and Calcium Imaging Methods in Wholemount Rat Retina
Published on: October 13, 2014
Detection of melanocortin-3 receptor mRNA in immature rat pituitary: functional relation to gamma3-MSH-induced
A Lorsignol1, V Vande Vijver, D Ramaekers
1Laboratory of Cell Pharmacology, University of Leuven, Medical School, Campus Gasthuisberg (O & N), Belgium.
Abstract:
We have previously shown that gamma3-MSH stimulates DNA replication in lactotrophs, somatotrophs and thyrotrophs of early postnatal rat pituitary in culture. Since the melanocortin-3 (MC-3) receptor is the only known receptor displaying high affinity for gamma3-MSH, the present study explored whether mRNA of the latter receptor is present in the pituitary and whether the receptor is functional. RT-PCR of RNA extracts from 14-day-old rat pituitary revealed the presence of MC-3 receptor mRNA in both the anterior and the neurointermediate lobe. The identity of the amplified products was confirmed by sequence analysis. Dispersed cells of 14-day-old female rats (24 h in culture) were exposed to gamma3-MSH, and changes in intracellular free calcium levels ([Ca2+]i) were assessed by means of fluo-3 video imaging. Gamma3-MSH evoked a rapid and maintained oscillating [Ca2+]i increase in 5%, 10% and 15% of the cells at a dose of 0.1, 1 and 10 nM, respectively. The MC-3/MC-4 receptor antagonist Ac-Nle4-c[Asp5,(D-Nal(2)7,Lys10]alpha-MSH-(4-10)-NH2 (SHU 9119) blocked the effect of gamma3-MSH in about 50% of the responsive cells. The present data suggest that the MC-3 receptor is expressed in the rat pituitary but that this receptor mediates only half of the effects of its putative ligand, gamma3-MSH, on [Ca2+]i. Part of the effect of gamma3-MSH may be mediated by a MC-3 receptor in a functional state different from the one studied previously in transfected cell lines or by a hitherto unidentified MC receptor.
Insights
Gamma3-melanocyte-stimulating hormone (gamma3-MSH) stimulates pituitary cell replication. This study confirms melanocortin-3 receptor (MC-3R) mRNA in rat pituitary and shows gamma3-MSH increases intracellular calcium, but only partially via MC-3R.
Area of Science:
- Endocrinology
- Molecular Biology
- Neuroscience
Background:
- Previous research demonstrated gamma3-melanocyte-stimulating hormone (gamma3-MSH) stimulates DNA replication in specific rat pituitary cell types.
- The melanocortin-3 receptor (MC-3R) is the primary known receptor for gamma3-MSH, suggesting a potential signaling pathway.
Purpose of the Study:
- To investigate the presence and functionality of the MC-3R in the rat pituitary gland.
- To determine if gamma3-MSH-induced intracellular calcium ([Ca2+]i) changes are mediated by the MC-3R.
Main Methods:
- Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect MC-3R mRNA in rat pituitary RNA extracts.
- Sequence analysis confirmed the identity of amplified MC-3R mRNA.
- Fluo-3 video imaging assessed changes in intracellular free calcium ([Ca2+]i) in cultured rat pituitary cells exposed to gamma3-MSH and an MC-3/MC-4 receptor antagonist (SHU 9119).
Main Results:
- MC-3R mRNA was detected in both the anterior and neurointermediate lobes of the 14-day-old rat pituitary.
- Gamma3-MSH induced a dose-dependent, oscillating increase in intracellular calcium ([Ca2+]i) in a subset of pituitary cells.
- The MC-3/MC-4 receptor antagonist SHU 9119 blocked approximately 50% of the gamma3-MSH-induced [Ca2+]i response.
Conclusions:
- The MC-3 receptor is expressed in the rat pituitary gland.
- While gamma3-MSH affects intracellular calcium levels in pituitary cells, the MC-3 receptor mediates only about half of this response.
- These findings suggest that either a distinct functional state of MC-3R or an unidentified melanocortin receptor contributes to gamma3-MSH signaling in the pituitary.
