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G-protein-independent coupling of MC4R to Kir7.1 in hypothalamic neurons.

Ghamari-Langroudi M, Digby GJ, Sebag JA, Millhauser GL, Palomino R, Matthews R, Gillyard T, Panaro BL, Tough IR, Cox HM, Denton JS, Cone RD - Nature (2015)

Bottom Line: Furthermore, AgRP is a biased agonist that hyperpolarizes neurons by binding to MC4R and opening Kir7.1, independently of its inhibition of α-MSH binding.Consequently, Kir7.1 signalling appears to be central to melanocortin-mediated regulation of energy homeostasis within the PVN.Coupling of MC4R to Kir7.1 may explain unusual aspects of the control of energy homeostasis by melanocortin signalling, including the gene dosage effect of MC4R and the sustained effects of AgRP on food intake.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Physiology &Biophysics, Vanderbilt University Medical Center, Nashville, Tennessee 37232, USA.

ABSTRACT
The regulated release of anorexigenic α-melanocyte stimulating hormone (α-MSH) and orexigenic Agouti-related protein (AgRP) from discrete hypothalamic arcuate neurons onto common target sites in the central nervous system has a fundamental role in the regulation of energy homeostasis. Both peptides bind with high affinity to the melanocortin-4 receptor (MC4R); existing data show that α-MSH is an agonist that couples the receptor to the Gαs signalling pathway, while AgRP binds competitively to block α-MSH binding and blocks the constitutive activity mediated by the ligand-mimetic amino-terminal domain of the receptor. Here we show that, in mice, regulation of firing activity of neurons from the paraventricular nucleus of the hypothalamus (PVN) by α-MSH and AgRP can be mediated independently of Gαs signalling by ligand-induced coupling of MC4R to closure of inwardly rectifying potassium channel, Kir7.1. Furthermore, AgRP is a biased agonist that hyperpolarizes neurons by binding to MC4R and opening Kir7.1, independently of its inhibition of α-MSH binding. Consequently, Kir7.1 signalling appears to be central to melanocortin-mediated regulation of energy homeostasis within the PVN. Coupling of MC4R to Kir7.1 may explain unusual aspects of the control of energy homeostasis by melanocortin signalling, including the gene dosage effect of MC4R and the sustained effects of AgRP on food intake.

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A model for α-MSH and AgRP signaling at PVN MC4R neuronsData presented here supports a model in which MC4R may couple to both Gαs signaling and regulation of Kir7.1 activity in PVN MC4R neurons. α-MSH results in elevation of intracellular cAMP through activation of Gαs, and inhibition of K+ efflux through Kir7.1, both of which are depolarizing. AgRP lowers the constitutive activity of the MC4R and blocks α-MSH binding, but data here shows that AgRP also acts as an agonist to increase K+ efflux through Kir7.1, producing a strong hyperpolarizing signal. The relative distribution and composition of the MC4R signaling complex in different subcellular compartments of PVN MC4R neurons has not been directly determined. Earlier models of α-MSH and AgRP action suggested competitive binding of these peptides to individual MC4R sites (orange box). Existing neuroanatomical data characterizing POMC and AgRP neuronal projections shows that α-MSH may act independently of AgRP at many sites in the CNS, since AgRP immunoreactive fibers are only observed in a subset of MC4R expressing nuclei containing POMC-immunoreative fibers (right circle, for review see 25). The ability of AgRP to act independently of α-MSH as a potent hyperpolarizing agonist, via regulation of Kir7.1, suggests the likely existence of independent AgRP sites of action (left circle). Recent reconstruction of EM images of the PVN in which POMC and AgRP containing synaptic vesicles have been specifically labeled with a genetically encoded marker provides preliminary anatomical support for this new model26. This study demonstrates that 52% of AgRP boutons in the PVN are not found in synapses, potentially supporting volume transmission of AgRP that may lead to competition with α-MSH at synaptic and/or non-synaptic sites. Additionally, the study found the vast majority of AgRP and POMC synaptic sites localized to different subcellular compartments of PVN neurons, supporting the independent action of both peptides. Synaptic release sites on soma were almost exclusively AgRP-containing, while POMC release sites were concentrated on distal dendrites. Another MC4R signaling pathway, involving cAMP/PKA-dependent activation of KATP channels and α-MSH induced hyperpolarization, has been demonstrated in MC4R neurons in the dorsal motor nucleus of the vagus in the brainstem (bottom right)21. Thus, while Kir7.1 signaling appears to be essential for depolarization of PVN MC4R neurons by α-MSH, Gαs signaling and elevation of cAMP may be depolarizing or hyperpolarizing, depending on the cellular context.
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Figure 14: A model for α-MSH and AgRP signaling at PVN MC4R neuronsData presented here supports a model in which MC4R may couple to both Gαs signaling and regulation of Kir7.1 activity in PVN MC4R neurons. α-MSH results in elevation of intracellular cAMP through activation of Gαs, and inhibition of K+ efflux through Kir7.1, both of which are depolarizing. AgRP lowers the constitutive activity of the MC4R and blocks α-MSH binding, but data here shows that AgRP also acts as an agonist to increase K+ efflux through Kir7.1, producing a strong hyperpolarizing signal. The relative distribution and composition of the MC4R signaling complex in different subcellular compartments of PVN MC4R neurons has not been directly determined. Earlier models of α-MSH and AgRP action suggested competitive binding of these peptides to individual MC4R sites (orange box). Existing neuroanatomical data characterizing POMC and AgRP neuronal projections shows that α-MSH may act independently of AgRP at many sites in the CNS, since AgRP immunoreactive fibers are only observed in a subset of MC4R expressing nuclei containing POMC-immunoreative fibers (right circle, for review see 25). The ability of AgRP to act independently of α-MSH as a potent hyperpolarizing agonist, via regulation of Kir7.1, suggests the likely existence of independent AgRP sites of action (left circle). Recent reconstruction of EM images of the PVN in which POMC and AgRP containing synaptic vesicles have been specifically labeled with a genetically encoded marker provides preliminary anatomical support for this new model26. This study demonstrates that 52% of AgRP boutons in the PVN are not found in synapses, potentially supporting volume transmission of AgRP that may lead to competition with α-MSH at synaptic and/or non-synaptic sites. Additionally, the study found the vast majority of AgRP and POMC synaptic sites localized to different subcellular compartments of PVN neurons, supporting the independent action of both peptides. Synaptic release sites on soma were almost exclusively AgRP-containing, while POMC release sites were concentrated on distal dendrites. Another MC4R signaling pathway, involving cAMP/PKA-dependent activation of KATP channels and α-MSH induced hyperpolarization, has been demonstrated in MC4R neurons in the dorsal motor nucleus of the vagus in the brainstem (bottom right)21. Thus, while Kir7.1 signaling appears to be essential for depolarization of PVN MC4R neurons by α-MSH, Gαs signaling and elevation of cAMP may be depolarizing or hyperpolarizing, depending on the cellular context.

Mentions: These data show the MC4R can depolarize or hyperpolarize hypothalamic PVN neurons in response to α-MSH or AgRP, respectively, through a novel G protein independent signaling pathway involving regulation of the activity of Kir7.1. While MC4R also is likely to couple to Gαs in most cells, cAMP/PKA-dependent activation of KATP channels, producing α-MSH induced hyperpolarization, has been demonstrated in MC4R neurons in the brainstem20. Thus, while Kir7.1 signaling appears to be essential for depolarization of PVN MC4R neurons by α-MSH, Gαs signaling and elevation of cAMP may be depolarizing or hyperpolarizing, depending on the cellular context (Extended Data Fig. 10).


G-protein-independent coupling of MC4R to Kir7.1 in hypothalamic neurons.

Ghamari-Langroudi M, Digby GJ, Sebag JA, Millhauser GL, Palomino R, Matthews R, Gillyard T, Panaro BL, Tough IR, Cox HM, Denton JS, Cone RD - Nature (2015)

A model for α-MSH and AgRP signaling at PVN MC4R neuronsData presented here supports a model in which MC4R may couple to both Gαs signaling and regulation of Kir7.1 activity in PVN MC4R neurons. α-MSH results in elevation of intracellular cAMP through activation of Gαs, and inhibition of K+ efflux through Kir7.1, both of which are depolarizing. AgRP lowers the constitutive activity of the MC4R and blocks α-MSH binding, but data here shows that AgRP also acts as an agonist to increase K+ efflux through Kir7.1, producing a strong hyperpolarizing signal. The relative distribution and composition of the MC4R signaling complex in different subcellular compartments of PVN MC4R neurons has not been directly determined. Earlier models of α-MSH and AgRP action suggested competitive binding of these peptides to individual MC4R sites (orange box). Existing neuroanatomical data characterizing POMC and AgRP neuronal projections shows that α-MSH may act independently of AgRP at many sites in the CNS, since AgRP immunoreactive fibers are only observed in a subset of MC4R expressing nuclei containing POMC-immunoreative fibers (right circle, for review see 25). The ability of AgRP to act independently of α-MSH as a potent hyperpolarizing agonist, via regulation of Kir7.1, suggests the likely existence of independent AgRP sites of action (left circle). Recent reconstruction of EM images of the PVN in which POMC and AgRP containing synaptic vesicles have been specifically labeled with a genetically encoded marker provides preliminary anatomical support for this new model26. This study demonstrates that 52% of AgRP boutons in the PVN are not found in synapses, potentially supporting volume transmission of AgRP that may lead to competition with α-MSH at synaptic and/or non-synaptic sites. Additionally, the study found the vast majority of AgRP and POMC synaptic sites localized to different subcellular compartments of PVN neurons, supporting the independent action of both peptides. Synaptic release sites on soma were almost exclusively AgRP-containing, while POMC release sites were concentrated on distal dendrites. Another MC4R signaling pathway, involving cAMP/PKA-dependent activation of KATP channels and α-MSH induced hyperpolarization, has been demonstrated in MC4R neurons in the dorsal motor nucleus of the vagus in the brainstem (bottom right)21. Thus, while Kir7.1 signaling appears to be essential for depolarization of PVN MC4R neurons by α-MSH, Gαs signaling and elevation of cAMP may be depolarizing or hyperpolarizing, depending on the cellular context.
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Figure 14: A model for α-MSH and AgRP signaling at PVN MC4R neuronsData presented here supports a model in which MC4R may couple to both Gαs signaling and regulation of Kir7.1 activity in PVN MC4R neurons. α-MSH results in elevation of intracellular cAMP through activation of Gαs, and inhibition of K+ efflux through Kir7.1, both of which are depolarizing. AgRP lowers the constitutive activity of the MC4R and blocks α-MSH binding, but data here shows that AgRP also acts as an agonist to increase K+ efflux through Kir7.1, producing a strong hyperpolarizing signal. The relative distribution and composition of the MC4R signaling complex in different subcellular compartments of PVN MC4R neurons has not been directly determined. Earlier models of α-MSH and AgRP action suggested competitive binding of these peptides to individual MC4R sites (orange box). Existing neuroanatomical data characterizing POMC and AgRP neuronal projections shows that α-MSH may act independently of AgRP at many sites in the CNS, since AgRP immunoreactive fibers are only observed in a subset of MC4R expressing nuclei containing POMC-immunoreative fibers (right circle, for review see 25). The ability of AgRP to act independently of α-MSH as a potent hyperpolarizing agonist, via regulation of Kir7.1, suggests the likely existence of independent AgRP sites of action (left circle). Recent reconstruction of EM images of the PVN in which POMC and AgRP containing synaptic vesicles have been specifically labeled with a genetically encoded marker provides preliminary anatomical support for this new model26. This study demonstrates that 52% of AgRP boutons in the PVN are not found in synapses, potentially supporting volume transmission of AgRP that may lead to competition with α-MSH at synaptic and/or non-synaptic sites. Additionally, the study found the vast majority of AgRP and POMC synaptic sites localized to different subcellular compartments of PVN neurons, supporting the independent action of both peptides. Synaptic release sites on soma were almost exclusively AgRP-containing, while POMC release sites were concentrated on distal dendrites. Another MC4R signaling pathway, involving cAMP/PKA-dependent activation of KATP channels and α-MSH induced hyperpolarization, has been demonstrated in MC4R neurons in the dorsal motor nucleus of the vagus in the brainstem (bottom right)21. Thus, while Kir7.1 signaling appears to be essential for depolarization of PVN MC4R neurons by α-MSH, Gαs signaling and elevation of cAMP may be depolarizing or hyperpolarizing, depending on the cellular context.
Mentions: These data show the MC4R can depolarize or hyperpolarize hypothalamic PVN neurons in response to α-MSH or AgRP, respectively, through a novel G protein independent signaling pathway involving regulation of the activity of Kir7.1. While MC4R also is likely to couple to Gαs in most cells, cAMP/PKA-dependent activation of KATP channels, producing α-MSH induced hyperpolarization, has been demonstrated in MC4R neurons in the brainstem20. Thus, while Kir7.1 signaling appears to be essential for depolarization of PVN MC4R neurons by α-MSH, Gαs signaling and elevation of cAMP may be depolarizing or hyperpolarizing, depending on the cellular context (Extended Data Fig. 10).

Bottom Line: Furthermore, AgRP is a biased agonist that hyperpolarizes neurons by binding to MC4R and opening Kir7.1, independently of its inhibition of α-MSH binding.Consequently, Kir7.1 signalling appears to be central to melanocortin-mediated regulation of energy homeostasis within the PVN.Coupling of MC4R to Kir7.1 may explain unusual aspects of the control of energy homeostasis by melanocortin signalling, including the gene dosage effect of MC4R and the sustained effects of AgRP on food intake.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Physiology &Biophysics, Vanderbilt University Medical Center, Nashville, Tennessee 37232, USA.

ABSTRACT
The regulated release of anorexigenic α-melanocyte stimulating hormone (α-MSH) and orexigenic Agouti-related protein (AgRP) from discrete hypothalamic arcuate neurons onto common target sites in the central nervous system has a fundamental role in the regulation of energy homeostasis. Both peptides bind with high affinity to the melanocortin-4 receptor (MC4R); existing data show that α-MSH is an agonist that couples the receptor to the Gαs signalling pathway, while AgRP binds competitively to block α-MSH binding and blocks the constitutive activity mediated by the ligand-mimetic amino-terminal domain of the receptor. Here we show that, in mice, regulation of firing activity of neurons from the paraventricular nucleus of the hypothalamus (PVN) by α-MSH and AgRP can be mediated independently of Gαs signalling by ligand-induced coupling of MC4R to closure of inwardly rectifying potassium channel, Kir7.1. Furthermore, AgRP is a biased agonist that hyperpolarizes neurons by binding to MC4R and opening Kir7.1, independently of its inhibition of α-MSH binding. Consequently, Kir7.1 signalling appears to be central to melanocortin-mediated regulation of energy homeostasis within the PVN. Coupling of MC4R to Kir7.1 may explain unusual aspects of the control of energy homeostasis by melanocortin signalling, including the gene dosage effect of MC4R and the sustained effects of AgRP on food intake.

Show MeSH
Related in: MedlinePlus