Expression of ghrelin receptor mRNA in the rat and the mouse brain

Abstract

Ghrelin is a hormone that stimulates growth hormone secretion and signals energy insufficiency via interaction with its receptor, the growth hormone secretagogue receptor (GHSR). The GHSR is located in both the central nervous system and the periphery. Its distribution in the CNS, as assessed by in situ hybridization histochemistry (ISHH), has been described previously in a few mammalian species, although these studies were limited by either the detail provided or the extent of the regions examined. In the present study, we systematically examined the distribution of GHSR mRNA in the adult rat and mouse brains and cervical spinal cords by using ISHH with novel cRNA probes specific for the mRNA encoding functional GHSR (the type 1a variant). We confirmed GHSR mRNA expression in several hypothalamic nuclei, many of which have long been recognized as playing roles in body weight and food intake. GHSR also was found in several other regions previously unknown to express GHSR mRNA, including many parasympathetic preganglionic neurons. Additionally, we found GHSR mRNA within all three components of the dorsal vagal complex, including the area postrema, the nucleus of the solitary tract, and the dorsal motor nucleus of the vagus. Finally, we examined the coexpression of GHSR with tyrosine hydroxylase and cholecystokinin and demonstrate a high degree of GHSR mRNA expression within dopaminergic, cholecystokinin-containing neurons of the substantia nigra and ventral tegmental area.

Figure 1

A series of low-power photomicrographs summarizing GHSR type 1a mRNA expression sites in the rat brain and cervical spinal cord. See Table 1 for an explanation of abbreviations. The scale bars represent 2 mm (and apply to all the panels in this figure).

Figure 1

A series of low-power photomicrographs summarizing GHSR type 1a mRNA expression sites in the rat brain and cervical spinal cord. See Table 1 for an explanation of abbreviations. The scale bars represent 2 mm (and apply to all the panels in this figure).

Figure 1

A series of low-power photomicrographs summarizing GHSR type 1a mRNA expression sites in the rat brain and cervical spinal cord. See Table 1 for an explanation of abbreviations. The scale bars represent 2 mm (and apply to all the panels in this figure).

Figure 2

A series of low-power photomicrographs summarizing GHSR type 1a mRNA expression sites in the mouse brain and cervical spinal cord. See Table 1 for an explanation of abbreviations. The scale bars represent 1 mm (and apply to all the panels in this figure).

Figure 3

A series of high-power photomicrographs demonstrating GHSR type 1a mRNA expression in the tuberal hypothalamus of the mouse. A, C, and E: rostrocaudally-arranged brain sections processed for GHSR mRNA by ISHH. B, D, and F: Nissl-stained brain sections adjacent to those appearing in A, C, and E. See Table 1 for an explanation of abbreviations. The scale bar located in panel E represents 200 μm (and applies to all the panels in this figure).

Figure 4

A series of photomicrographs demonstrating co-expression of GHSR and tyrosine hydroxylase in the rat arcuate nucleus and GHSR type 1a mRNA expression in the rat facial motor nucleus. A and D: rostrocaudally-arranged brain sections through the tuberal hypothalamus. TH-IR cell bodies are stained orange-brown. Cells expressing GHSR type 1a mRNA have overlying black silver granules. The boxed areas in A and D are magnified in B and E, and the boxed areas in B and E are magnified in C and F. G: facial motor nucleus. Cells expressing GHSR type 1a mRNA have overlying black silver granules. The orange-brown coloration indicates CCK-IR. The boxed area in G is magnified in H, and the boxed area in H is magnified in I. Black arrows indicate examples of neurons doubly-labeled with GHSR riboprobe and either anti-TH or anti-CCK antisera. Abbreviations: dc, dorsal cap subdivision of facial motor nucleus (7); i, intermediate subdivision of 7; l, lateral subdivision of 7; m, medial subdivision of 7. The scale bar located in panel D represents 200 μm (and applies to panels A and D); the scale bar located in panel E represents 50 μm (and applies to panels B and E); the scale bar located in panel F represents 20 μm (and applies to panels C and F); the scale bar located in panel G represents 50 μm; the scale bar located in panel H represents 25 μm; the scale bar located in panel I represents 10 μm.

Figure 5

A series of high-power photomicrographs demonstrating GHSR type 1a mRNA expression in the dorsal vagal complex of the rat. A, C, E, and G: rostrocaudally-arranged brain sections processed for GHSR mRNA by ISHH. B, D, F, and H: Nissl-stained brain sections adjacent to those appearing in A, C, E, and G. See Table 1 for an explanation of abbreviations. The scale bar located in panel G represents 200 μm (and applies to all the panels in this figure).

Figure 6

A series of high-power photomicrographs demonstrating GHSR type 1a mRNA expression in the dorsal vagal complex of the mouse. A and C: rostrocaudally-arranged brain sections processed for GHSR mRNA by ISHH. B and D: Nissl-stained brain sections adjacent to those appearing in A and B. See Table 1 for an explanation of abbreviations. The scale bar located in panel C represents 200 μm (and applies to all the panels in this figure).

Figure 7

A series of photomicrographs demonstrating co-expression of GHSR with tyrosine hydroxylase and with cholecystokinin in the rat midbrain. A – F: co-expression of GHSR and TH. A, B, and C: rostrocaudally-arranged brain sections through the midbrain. The boxed areas in A, B, and C are magnified in D, E, and F. D: SNC. E: SNL. F: VTA. TH-IR neurons are stained orange brown. Cells expressing GHSR type 1a mRNA have overlying black silver granules. G – L: co-expression of GHSR and CCK. The boxed areas in G, H, and I are magnified in J, K, and L. G: VTA and SNC. H: SNC and SNL. I: EW. J: VTA (upper left corner) and SNC (lower right corner). K: SNL. L: EW. CCK-IR neurons are stained orange-brown. Cells expressing GHSR type 1a mRNA have overlying black silver granules. See Table 1 for an explanation of abbreviations. Black arrows indicate examples of neurons doubly-labeled with GHSR riboprobe and either anti-TH or anti-CCK antisera. The scale bar located in panel C represents 500 μm (and applies to panels A – C); the scale bar located in panel F represents 20 μm (and applies to panels D – F); the scale bar located in panel I represents 100 μm (and applies to panels G – I); the scale bar located in panel L represents 20 μm (and applies to panels J – L).

Figure 8

A series of photomicrographs demonstrating distinct expression patterns for GHSR and cholecystokinin in the dorsal vagal complex of the rat. A and C: rostrocaudally-arranged brain sections through the dorsal vagal complex. CCK-IR cell bodies are stained orange-brown. Cells expressing GHSR type 1a mRNA have overlying black silver granules. The boxed areas in A and C are magnified in B and D. Black arrows indicate examples of neurons singly-labeled with GHSR riboprobe. White arrows indicate examples of neurons singly-labeled with anti-CCK antiserum. The scale bar located in panel C represents 200 μm (and applies to panels A and C); the scale bar located in panel D represents 50 μm (and applies to panels B and D).