Neuronal M ₃ muscarinic acetylcholine receptors are essential for somatotroph proliferation and normal somatic growth

Abstract

The molecular pathways that promote the proliferation and maintenance of pituitary somatotrophs and other cell types of the anterior pituitary gland are not well understood at present. However, such knowledge is likely to lead to the development of novel drugs useful for the treatment of various human growth disorders. Although muscarinic cholinergic pathways have been implicated in regulating somatotroph function, the physiological relevance of this effect and the localization and nature of the receptor subtypes involved in this activity remain unclear. We report the surprising observation that mutant mice that selectively lack the M3 muscarinic acetylcholine receptor subtype in the brain (neurons and glial cells; Br-M3-KO mice) showed a dwarf phenotype associated with a pronounced hypoplasia of the anterior pituitary gland and a marked decrease in pituitary and serum growth hormone (GH) and prolactin. anterior pituitary gland ͉ dwarfism ͉ knockout mice V arious neurotransmitter systems have been implicated in regulating somatotroph function and growth hormone (GH) secretion, either by acting on the anterior pituitary gland directly or by modulating the release of GH-releasing hormone (GHRH) or somatostatin from the hypothalamus (for a comprehensive review, see ref. 1). However, the physiological relevance of the individual pathways and the identity and localization of the receptor subtypes involved in mediating these effects are not well understood in most cases.Pharmacological studies suggest that central muscarinic cholinergic pathways play a role in stimulating GH release in experimental animals and humans (1-9). The identification of the molecular pathways and the specific muscarinic ACh receptor (mAChR) subtypes involved in mediating these responses should be of considerable potential therapeutic interest. However, work in this area has been complicated by the existence of 5 molecularly distinct mAChR subtypes (M 1 -M 5 ) that are difficult to distinguish by classical pharmacological tools (10, 11). Moreover, virtually all brain regions express multiple mAChRs in a complex, overlapping pattern (12-15).The M 3 mAChR subtype is expressed at relatively high levels in the hypothalamus but is also found in many other brain regions (16-18). At present, little is known about the physiological relevance of these neuronal M 3 mAChRs. To shed light on this issue, we used Cre/loxP technology to generate a set of mutant mice, referred to as Br-M3-KO mice, that selectively lacked the M 3 mAChR subtype in the brain (neurons and glial cells). Strikingly, Br-M3-KO mice showed a dwarf phenotype, associated with a dramatic reduction in the size of the anterior pituitary gland, a marked decrease in pituitary GH and prolactin levels, and significantly reduced serum GH, insulin-like growth factor-1 (IGF-1), and prolactin levels. Remarkably, treatment of Br-M3-KO mice with CJC-1295, a synthetic GHRH analog (19-21), restored normal pituitary size, serum GH and IGF-1 levels, and longitudinal growth. These data reveal an unexpect...

Structural Basis of M3 Muscarinic Receptor Dimer/Oligomer Formation

“…The M3R, a prototypic class A GPCR that preferentially couples to G proteins of the G q family, is involved in numerous important physiological functions (34 -36). For example, recent studies with M3R mutant mice have shown that M3Rs expressed by pancreatic ␤ cells play a key role in maintaining normal blood glucose levels (37) and that neuronal M3Rs are required for proper somatic growth (38) and bone formation (39). Moreover, Raufman et al (40) recently demonstrated that enhanced M3R activity may play a role in the pathogenesis of colon cancer.…”

Novel Muscarinic Receptor Mutant Mouse Models

“…These observations suggest that central M 3 receptors do not play a significant role in regulating these processes. Interestingly, however, the brain-M3-KO mice displayed a dwarf-like appearance (adult mutant mice were~10% shorter than control littermates; Gautam et al 2009). This phenotype was associated with a significant reduction in the serum levels of growth hormone (GH) and insulin-like growth factor I (IGF-1; Fig.…”

“…To shed light onto the roles of central M 3 mAChRs, Gautam et al (2009) used Cre/loxP technology to generate mutant mice that lacked M 3 receptors specifically in neurons and glial cells (brain-M3-KO or Br-M3-KO mice). These mice were obtained by crossing a Cre transgene driven by the nestin promoter into mice that were homozygous for a floxed version of the M 3 receptor gene.…”

“…**p < 0.01, as compared to the corresponding control group. Data were taken from Gautam et al (2009) Current evidence suggests that IGF-1, following its GH-dependent release from the liver, is the major factor mediating the stimulatory effect of GH on longitudinal growth. It is therefore likely that the reduction in body length displayed by the Br-M3-KO mice is the direct consequence of decreased GH and IGF-1 levels.…”

“…Since the anterior pituitary is not of neuronal origin, the nestin-Cre transgene is not expressed in this part of the pituitary (Tronche et al 1999;Wettschureck et al 2005). Gautam et al (2009) therefore speculated that the primary defect leading to the hypoplasia of the anterior pituitary in the Br-M3-KO mice resides outside of the pituitary itself. Consistent with this notion, the authors found that hypothalamic GHRH neurons express M 3 mAChRs and that hypothalamic GHRH levels were greatly reduced in Br-M3-KO mice.…”