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Critical role of neuropeptides B/W receptor 1 signaling in social behavior and fear memory.

Nagata-Kuroiwa R, Furutani N, Hara J, Hondo M, Ishii M, Abe T, Mieda M, Tsujino N, Motoike T, Yanagawa Y, Kuwaki T, Yamamoto M, Yanagisawa M, Sakurai T - PLoS ONE (2011)

Bottom Line: These data suggest that NPBWR1 plays a critical role in limbic system function and stress responses.Histological and electrophysiological studies showed that NPBWR1 acts as an inhibitory regulator on a subpopulation of GABAergic neurons in the lateral division of the CeA and terminates stress responses.These findings suggest important roles of NPBWR1 in regulating amygdala function during physical and social stress.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Neuroscience and Integrative Physiology, Faculty of Medicine, Kanazawa University, Kanazawa, Japan.

ABSTRACT
Neuropeptide B/W receptor 1 (NPBWR1) is a G-protein coupled receptor, which was initially reported as an orphan receptor, and whose ligands were identified by this and other groups in 2002 and 2003. To examine the physiological roles of NPBWR1, we examined phenotype of Npbwr1⁻/⁻ mice. When presented with an intruder mouse, Npbwr1⁻/⁻ mice showed impulsive contact with the strange mice, produced more intense approaches toward them, and had longer contact and chasing time along with greater and sustained elevation of heart rate and blood pressure compared to wild type mice. Npbwr1⁻/⁻ mice also showed increased autonomic and neuroendocrine responses to physical stress, suggesting that impairment of NPBWR1 leads to stress vulnerability. We also observed that these mice show abnormality in the contextual fear conditioning test. These data suggest that NPBWR1 plays a critical role in limbic system function and stress responses. Histological and electrophysiological studies showed that NPBWR1 acts as an inhibitory regulator on a subpopulation of GABAergic neurons in the lateral division of the CeA and terminates stress responses. These findings suggest important roles of NPBWR1 in regulating amygdala function during physical and social stress.

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Function of NPBWR1 in regulation of CeA neurons.(A) Left panel, Dual-label In situ hybridization histochemistry showed co-localization of Npbwr1 mRNA (blue) with Gad67-expressing neurons (red) in the CeAl of mice. Scale bar equals 250 µm. Middle panel, higher power view of yellow rectangle region in the left panel. Right panel, high power view of yellow rectangle region in the middle panel. Opt, optic tract. (B) Left panels, typical examples of whole cell patch-clamp recording from GAD67-expressing neurons in Gad67-gfp brain sections, showing that bath-application of NPB (upper panel, 500 nM) or NPW (lower panel, 500 nM) potently inhibited neuronal activity. Right panel, numbers of GFP-positive neurons activated or inhibited by NPB/W application. We did not observe any effects in neurons of Npbwr1−/− mice. (C) A typical example of morphology of NPB/W-inhibited GABAergic neurons as revealed by neurobiotin injection after patch-clamp recordings. This cell resides in the medial region of the CeAl and sends long projections to outside of the amygdala. (D) Schematic drawings of axonal projections of NPW and/or NPB-inhibited neurons in the CeAl. Left panel shows three neurons depicted in different colors that send axons within the CeAl. Right panel shows four neurons that send axons outside of the CeA.
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pone-0016972-g004: Function of NPBWR1 in regulation of CeA neurons.(A) Left panel, Dual-label In situ hybridization histochemistry showed co-localization of Npbwr1 mRNA (blue) with Gad67-expressing neurons (red) in the CeAl of mice. Scale bar equals 250 µm. Middle panel, higher power view of yellow rectangle region in the left panel. Right panel, high power view of yellow rectangle region in the middle panel. Opt, optic tract. (B) Left panels, typical examples of whole cell patch-clamp recording from GAD67-expressing neurons in Gad67-gfp brain sections, showing that bath-application of NPB (upper panel, 500 nM) or NPW (lower panel, 500 nM) potently inhibited neuronal activity. Right panel, numbers of GFP-positive neurons activated or inhibited by NPB/W application. We did not observe any effects in neurons of Npbwr1−/− mice. (C) A typical example of morphology of NPB/W-inhibited GABAergic neurons as revealed by neurobiotin injection after patch-clamp recordings. This cell resides in the medial region of the CeAl and sends long projections to outside of the amygdala. (D) Schematic drawings of axonal projections of NPW and/or NPB-inhibited neurons in the CeAl. Left panel shows three neurons depicted in different colors that send axons within the CeAl. Right panel shows four neurons that send axons outside of the CeA.

Mentions: Since we found abnormality of social interaction, autonomic responses, and contextual fear conditioning, all of which are related to amygdala function, in Npbwr1−/− mice, we next explored the neuronal mechanisms by which NPBWR1 regulates the function of the amygdala, by probing the expression profile of NPBWR1 in the neural circuitry of the amygdala in mice. By double-label in situ hybridization, we found that Npbwr1 was abundantly expressed in GAD67-positive, gamma-aminobutyric acid (GABAergic) neurons in the medial region of the lateral division of the CeA (CeAl) (Fig. 4A). Npbwr1 mRNA was present in 34.1±5.3% (n = 3) of Gad67-positive neurons within the CeAl. Virtually all Npbwr1-positive neurons were also positive for Gad67, suggesting that most of the NPBWR1-positive neurons were GABAergic in the CeAl. We also observed that Npbwr1 was expressed in Gad67-positive neurons in the BST, which is recognized to be an extension of the CeA [9] (Fig. S2A). These findings confirm that NPBWR1 is expressed in GABAergic neurons in the output nuclei of the extended amygdala, where NPW-immunoreactive fibers were exclusively observed in the mouse brain [5], [7] (Fig. S2B). We next examined the effect of NPB and NPW on Gad67-positive neurons in the CeAl by means of patch-clamp recording. Whole cell recording showed that bath application of NPB or NPW hyperpolarized and inhibited 8 out of 19 Gad67-positive neurons in the CeAl in slice preparations (Fig. 4B). None of the 10 neurons tested from Npbwr1−/− mice showed such inhibition. Neurons in the CeAl are mostly GABAergic and many of these neurons are thought to send inhibitory projections to neurons in the medial part of CeA (CeAm), the main output nucleus of the amygdala. However, a subpopulation of CeAl neurons are also known to directly project to the BST and brain stem target areas [14]. Morphological examination of NPW-inhibited cells by injecting neurobiotin after recordings showed that four out of seven NPW-inhibited neurons examined had relatively long axons that projected through the CeAm to outside the amygdala (Fig. 4C, D). These observations demonstrate that NPB/W acts on projection neurons in the CeAl. We also observed some cells with shorter axons that ended within the CeAl. Because these studies were done using slice preparations, we cannot conclude that these axons ended within the CeAl, but it is plausible that some NPBWR1-positive neurons could be GABAergic interneurons in the CeAl.


Critical role of neuropeptides B/W receptor 1 signaling in social behavior and fear memory.

Nagata-Kuroiwa R, Furutani N, Hara J, Hondo M, Ishii M, Abe T, Mieda M, Tsujino N, Motoike T, Yanagawa Y, Kuwaki T, Yamamoto M, Yanagisawa M, Sakurai T - PLoS ONE (2011)

Function of NPBWR1 in regulation of CeA neurons.(A) Left panel, Dual-label In situ hybridization histochemistry showed co-localization of Npbwr1 mRNA (blue) with Gad67-expressing neurons (red) in the CeAl of mice. Scale bar equals 250 µm. Middle panel, higher power view of yellow rectangle region in the left panel. Right panel, high power view of yellow rectangle region in the middle panel. Opt, optic tract. (B) Left panels, typical examples of whole cell patch-clamp recording from GAD67-expressing neurons in Gad67-gfp brain sections, showing that bath-application of NPB (upper panel, 500 nM) or NPW (lower panel, 500 nM) potently inhibited neuronal activity. Right panel, numbers of GFP-positive neurons activated or inhibited by NPB/W application. We did not observe any effects in neurons of Npbwr1−/− mice. (C) A typical example of morphology of NPB/W-inhibited GABAergic neurons as revealed by neurobiotin injection after patch-clamp recordings. This cell resides in the medial region of the CeAl and sends long projections to outside of the amygdala. (D) Schematic drawings of axonal projections of NPW and/or NPB-inhibited neurons in the CeAl. Left panel shows three neurons depicted in different colors that send axons within the CeAl. Right panel shows four neurons that send axons outside of the CeA.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3044739&req=5

pone-0016972-g004: Function of NPBWR1 in regulation of CeA neurons.(A) Left panel, Dual-label In situ hybridization histochemistry showed co-localization of Npbwr1 mRNA (blue) with Gad67-expressing neurons (red) in the CeAl of mice. Scale bar equals 250 µm. Middle panel, higher power view of yellow rectangle region in the left panel. Right panel, high power view of yellow rectangle region in the middle panel. Opt, optic tract. (B) Left panels, typical examples of whole cell patch-clamp recording from GAD67-expressing neurons in Gad67-gfp brain sections, showing that bath-application of NPB (upper panel, 500 nM) or NPW (lower panel, 500 nM) potently inhibited neuronal activity. Right panel, numbers of GFP-positive neurons activated or inhibited by NPB/W application. We did not observe any effects in neurons of Npbwr1−/− mice. (C) A typical example of morphology of NPB/W-inhibited GABAergic neurons as revealed by neurobiotin injection after patch-clamp recordings. This cell resides in the medial region of the CeAl and sends long projections to outside of the amygdala. (D) Schematic drawings of axonal projections of NPW and/or NPB-inhibited neurons in the CeAl. Left panel shows three neurons depicted in different colors that send axons within the CeAl. Right panel shows four neurons that send axons outside of the CeA.
Mentions: Since we found abnormality of social interaction, autonomic responses, and contextual fear conditioning, all of which are related to amygdala function, in Npbwr1−/− mice, we next explored the neuronal mechanisms by which NPBWR1 regulates the function of the amygdala, by probing the expression profile of NPBWR1 in the neural circuitry of the amygdala in mice. By double-label in situ hybridization, we found that Npbwr1 was abundantly expressed in GAD67-positive, gamma-aminobutyric acid (GABAergic) neurons in the medial region of the lateral division of the CeA (CeAl) (Fig. 4A). Npbwr1 mRNA was present in 34.1±5.3% (n = 3) of Gad67-positive neurons within the CeAl. Virtually all Npbwr1-positive neurons were also positive for Gad67, suggesting that most of the NPBWR1-positive neurons were GABAergic in the CeAl. We also observed that Npbwr1 was expressed in Gad67-positive neurons in the BST, which is recognized to be an extension of the CeA [9] (Fig. S2A). These findings confirm that NPBWR1 is expressed in GABAergic neurons in the output nuclei of the extended amygdala, where NPW-immunoreactive fibers were exclusively observed in the mouse brain [5], [7] (Fig. S2B). We next examined the effect of NPB and NPW on Gad67-positive neurons in the CeAl by means of patch-clamp recording. Whole cell recording showed that bath application of NPB or NPW hyperpolarized and inhibited 8 out of 19 Gad67-positive neurons in the CeAl in slice preparations (Fig. 4B). None of the 10 neurons tested from Npbwr1−/− mice showed such inhibition. Neurons in the CeAl are mostly GABAergic and many of these neurons are thought to send inhibitory projections to neurons in the medial part of CeA (CeAm), the main output nucleus of the amygdala. However, a subpopulation of CeAl neurons are also known to directly project to the BST and brain stem target areas [14]. Morphological examination of NPW-inhibited cells by injecting neurobiotin after recordings showed that four out of seven NPW-inhibited neurons examined had relatively long axons that projected through the CeAm to outside the amygdala (Fig. 4C, D). These observations demonstrate that NPB/W acts on projection neurons in the CeAl. We also observed some cells with shorter axons that ended within the CeAl. Because these studies were done using slice preparations, we cannot conclude that these axons ended within the CeAl, but it is plausible that some NPBWR1-positive neurons could be GABAergic interneurons in the CeAl.

Bottom Line: These data suggest that NPBWR1 plays a critical role in limbic system function and stress responses.Histological and electrophysiological studies showed that NPBWR1 acts as an inhibitory regulator on a subpopulation of GABAergic neurons in the lateral division of the CeA and terminates stress responses.These findings suggest important roles of NPBWR1 in regulating amygdala function during physical and social stress.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Neuroscience and Integrative Physiology, Faculty of Medicine, Kanazawa University, Kanazawa, Japan.

ABSTRACT
Neuropeptide B/W receptor 1 (NPBWR1) is a G-protein coupled receptor, which was initially reported as an orphan receptor, and whose ligands were identified by this and other groups in 2002 and 2003. To examine the physiological roles of NPBWR1, we examined phenotype of Npbwr1⁻/⁻ mice. When presented with an intruder mouse, Npbwr1⁻/⁻ mice showed impulsive contact with the strange mice, produced more intense approaches toward them, and had longer contact and chasing time along with greater and sustained elevation of heart rate and blood pressure compared to wild type mice. Npbwr1⁻/⁻ mice also showed increased autonomic and neuroendocrine responses to physical stress, suggesting that impairment of NPBWR1 leads to stress vulnerability. We also observed that these mice show abnormality in the contextual fear conditioning test. These data suggest that NPBWR1 plays a critical role in limbic system function and stress responses. Histological and electrophysiological studies showed that NPBWR1 acts as an inhibitory regulator on a subpopulation of GABAergic neurons in the lateral division of the CeA and terminates stress responses. These findings suggest important roles of NPBWR1 in regulating amygdala function during physical and social stress.

Show MeSH
Related in: MedlinePlus