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GABA Signaling and Neuroactive Steroids in Adrenal Medullary Chromaffin Cells.

Harada K, Matsuoka H, Fujihara H, Ueta Y, Yanagawa Y, Inoue M - Front Cell Neurosci (2016)

Bottom Line: GABA has two actions mediated by GABAA receptors in chromaffin cells: it induces catecholamine secretion by itself and produces an inhibition of synaptically evoked secretion by a shunt effect.This function of GABA may be facilitated by expression of the immature isoforms of GAD and GABAA receptors and the lack of expression of plasma membrane GABA transporters (GATs).In this review, we will consider how the para/autocrine function of GABA is achieved, focusing on the structural and molecular mechanisms for GABA signaling.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Systems Physiology, University of Occupational and Environmental Health School of Medicine Kitakyushu, Japan.

ABSTRACT
Gamma-aminobutyric acid (GABA) is produced not only in the brain, but also in endocrine cells by the two isoforms of glutamic acid decarboxylase (GAD), GAD65 and GAD67. In rat adrenal medullary chromaffin cells only GAD67 is expressed, and GABA is stored in large dense core vesicles (LDCVs), but not synaptic-like microvesicles (SLMVs). The α3β2/3γ2 complex represents the majority of GABAA receptors expressed in rat and guinea pig chromaffin cells, whereas PC12 cells, an immortalized rat chromaffin cell line, express the α1 subunit as well as the α3. The expression of α3, but not α1, in PC12 cells is enhanced by glucocorticoid activity, which may be mediated by both the mineralocorticoid receptor (MR) and the glucocorticoid receptor (GR). GABA has two actions mediated by GABAA receptors in chromaffin cells: it induces catecholamine secretion by itself and produces an inhibition of synaptically evoked secretion by a shunt effect. Allopregnanolone, a neuroactive steroid which is secreted from the adrenal cortex, produces a marked facilitation of GABAA receptor channel activity. Since there are no GABAergic nerve fibers in the adrenal medulla, GABA may function as a para/autocrine factor in the chromaffin cells. This function of GABA may be facilitated by expression of the immature isoforms of GAD and GABAA receptors and the lack of expression of plasma membrane GABA transporters (GATs). In this review, we will consider how the para/autocrine function of GABA is achieved, focusing on the structural and molecular mechanisms for GABA signaling.

No MeSH data available.


Related in: MedlinePlus

Enhancement of expression of the GABAA α3 subunit in PC12 cells by dexamethasone. (A,B) Immunofluorescent staining for α1 and α3 subunits in PC12 cells, respectively. Cells were cultured for 1 week in Dulbecco’s modified medium supplemented with 10% fetal bovine serum (FBS; control), Dulbecco’s modified medium with 10 μM dexamethasone (+DM), or Dulbecco’s modified medium (−FBS). Left and right images represent confocal fluorescent images and merge of fluorescent and DIC images. (C,D) Summaries of α1- and α3-like immunoreactivities, respectively. The amounts of α1- and α3-like immunoreactivities were measured by using ImageJ software (NIH, Bethesda, MD, USA). The amounts of the immunoreactivities in PC12 cells cultured with DM (+DM) and without DM and FBS (−FBS) were expressed as percentages of those in the control PC12 cells, which were measured under the same conditions. The data indicate mean ± SEM (C, n = 12, 12, and 10 for control, +DM, and −FBS, respectively; D, n = 19, 9, and 10 for control, +DM, and −FBS, respectively). #and ##represent statistical significance of P < 0.01 and P < 0.001, respectively. (B,D) Are reproduced from Inoue et al. (2013).
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Figure 2: Enhancement of expression of the GABAA α3 subunit in PC12 cells by dexamethasone. (A,B) Immunofluorescent staining for α1 and α3 subunits in PC12 cells, respectively. Cells were cultured for 1 week in Dulbecco’s modified medium supplemented with 10% fetal bovine serum (FBS; control), Dulbecco’s modified medium with 10 μM dexamethasone (+DM), or Dulbecco’s modified medium (−FBS). Left and right images represent confocal fluorescent images and merge of fluorescent and DIC images. (C,D) Summaries of α1- and α3-like immunoreactivities, respectively. The amounts of α1- and α3-like immunoreactivities were measured by using ImageJ software (NIH, Bethesda, MD, USA). The amounts of the immunoreactivities in PC12 cells cultured with DM (+DM) and without DM and FBS (−FBS) were expressed as percentages of those in the control PC12 cells, which were measured under the same conditions. The data indicate mean ± SEM (C, n = 12, 12, and 10 for control, +DM, and −FBS, respectively; D, n = 19, 9, and 10 for control, +DM, and −FBS, respectively). #and ##represent statistical significance of P < 0.01 and P < 0.001, respectively. (B,D) Are reproduced from Inoue et al. (2013).

Mentions: The fact that the α3β2/3γ2 complex constitutes the majority of GABAA receptors means chromaffin cells continue to express immature GABAA receptors in adulthood, which raises a question about how the control of subunit expression may differ between adrenal gland and brain. The internal environment drastically changes in the growth process from the prenatal period to the neonate and then juvenile periods. One such change in rats is a decrease in blood glucocorticoid during the first 2 weeks of life (Sapolsky and Meaney, 1986). Early life experience, including impaired maternal care, influences brain development and the expression of GABAA receptors in the adulthood (Caldji et al., 1998, 2003), and the blood glucocorticoid concentration increases in rats subjected to maternal deprivation (Chen et al., 2012). These observations suggest that glucocorticoids play an important role for regulation of GABAA receptors. To examine the effects of glucocorticoids on the expression of GABAA receptors, PC12 cells were used. PC12 cells were found to express α1 as well as α3 subunits, which were mainly present at the cell periphery (Inoue et al., 2013). The expression of both subunits was abolished by removal of serum from the culture medium, but that of α3—but not α1—was restored by addition of the synthetic glucocorticoid dexamethasone (Figure 2), indicating that glucocorticoid activity supports α3 expression. There are two kinds of receptors in the cytosol to which glucocorticoids bind: one is mineralocorticoid receptor (MR); the other is glucocorticoid receptor (GR; de Kloet et al., 2005). The GR and MR function either as a homodimer or heterodimer, and the affinity of MR for cortical steroids is 10-fold higher than that of GR (de Kloet et al., 2005). The expression of α3, but not α1, in PC12 cells was suppressed in a dose-dependent manner by not only mifepristone (Inoue et al., 2013), a specific GR inhibitor (Reul et al., 1990), but also by RU28318 (unpublished observations by KH and MI), a specific MR inhibitor (Ulmann et al., 1985), suggesting the involvement of MR as well as GR. In fact, the expression of MR in PC12 cells has been reported (Goto et al., 2009). Whether MR is involved in the regulation of α3 expression in adrenal chromaffin cells or not remains to be explored. One critical difference between adrenal medulla and brain may be the existence of a higher glucocorticoid level in the extracellular space of adrenal medulla (Wurtman, 2002). Thus, based on the fingdings in PC12 cells, glucocorticoids likely preserve α3 expression in adrenal chromaffin cells. Recently, a decrease in intracellular Cl− concentration ([Cl−]i) has been shown to be involved in the substitution of α1 for α3 (Succol et al., 2012). The value of [Cl−]i in neurons is determined by a balance between Na+, K+, Cl−-cotransporter type 1 (NKCC1) and K+, Cl−-cotransporter type 2 (KCC2), which are responsible for import and export of Cl−, respectively (Kaila et al., 2014). Aldosterone has been reported to enhance NKCC1 activity in vascular smooth muscle cells (Ding et al., 2014) with the consequent increase in [Cl−]i (Davis et al., 1993). These results raise the possibility that glucocorticoids enhance NKCC1 activity in chromaffin cells, thereby preserving α3-containing GABAA receptors. A further study will be required to elucidate the molecular mechanism for α3 preservation in chromaffin cells.


GABA Signaling and Neuroactive Steroids in Adrenal Medullary Chromaffin Cells.

Harada K, Matsuoka H, Fujihara H, Ueta Y, Yanagawa Y, Inoue M - Front Cell Neurosci (2016)

Enhancement of expression of the GABAA α3 subunit in PC12 cells by dexamethasone. (A,B) Immunofluorescent staining for α1 and α3 subunits in PC12 cells, respectively. Cells were cultured for 1 week in Dulbecco’s modified medium supplemented with 10% fetal bovine serum (FBS; control), Dulbecco’s modified medium with 10 μM dexamethasone (+DM), or Dulbecco’s modified medium (−FBS). Left and right images represent confocal fluorescent images and merge of fluorescent and DIC images. (C,D) Summaries of α1- and α3-like immunoreactivities, respectively. The amounts of α1- and α3-like immunoreactivities were measured by using ImageJ software (NIH, Bethesda, MD, USA). The amounts of the immunoreactivities in PC12 cells cultured with DM (+DM) and without DM and FBS (−FBS) were expressed as percentages of those in the control PC12 cells, which were measured under the same conditions. The data indicate mean ± SEM (C, n = 12, 12, and 10 for control, +DM, and −FBS, respectively; D, n = 19, 9, and 10 for control, +DM, and −FBS, respectively). #and ##represent statistical significance of P < 0.01 and P < 0.001, respectively. (B,D) Are reproduced from Inoue et al. (2013).
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Figure 2: Enhancement of expression of the GABAA α3 subunit in PC12 cells by dexamethasone. (A,B) Immunofluorescent staining for α1 and α3 subunits in PC12 cells, respectively. Cells were cultured for 1 week in Dulbecco’s modified medium supplemented with 10% fetal bovine serum (FBS; control), Dulbecco’s modified medium with 10 μM dexamethasone (+DM), or Dulbecco’s modified medium (−FBS). Left and right images represent confocal fluorescent images and merge of fluorescent and DIC images. (C,D) Summaries of α1- and α3-like immunoreactivities, respectively. The amounts of α1- and α3-like immunoreactivities were measured by using ImageJ software (NIH, Bethesda, MD, USA). The amounts of the immunoreactivities in PC12 cells cultured with DM (+DM) and without DM and FBS (−FBS) were expressed as percentages of those in the control PC12 cells, which were measured under the same conditions. The data indicate mean ± SEM (C, n = 12, 12, and 10 for control, +DM, and −FBS, respectively; D, n = 19, 9, and 10 for control, +DM, and −FBS, respectively). #and ##represent statistical significance of P < 0.01 and P < 0.001, respectively. (B,D) Are reproduced from Inoue et al. (2013).
Mentions: The fact that the α3β2/3γ2 complex constitutes the majority of GABAA receptors means chromaffin cells continue to express immature GABAA receptors in adulthood, which raises a question about how the control of subunit expression may differ between adrenal gland and brain. The internal environment drastically changes in the growth process from the prenatal period to the neonate and then juvenile periods. One such change in rats is a decrease in blood glucocorticoid during the first 2 weeks of life (Sapolsky and Meaney, 1986). Early life experience, including impaired maternal care, influences brain development and the expression of GABAA receptors in the adulthood (Caldji et al., 1998, 2003), and the blood glucocorticoid concentration increases in rats subjected to maternal deprivation (Chen et al., 2012). These observations suggest that glucocorticoids play an important role for regulation of GABAA receptors. To examine the effects of glucocorticoids on the expression of GABAA receptors, PC12 cells were used. PC12 cells were found to express α1 as well as α3 subunits, which were mainly present at the cell periphery (Inoue et al., 2013). The expression of both subunits was abolished by removal of serum from the culture medium, but that of α3—but not α1—was restored by addition of the synthetic glucocorticoid dexamethasone (Figure 2), indicating that glucocorticoid activity supports α3 expression. There are two kinds of receptors in the cytosol to which glucocorticoids bind: one is mineralocorticoid receptor (MR); the other is glucocorticoid receptor (GR; de Kloet et al., 2005). The GR and MR function either as a homodimer or heterodimer, and the affinity of MR for cortical steroids is 10-fold higher than that of GR (de Kloet et al., 2005). The expression of α3, but not α1, in PC12 cells was suppressed in a dose-dependent manner by not only mifepristone (Inoue et al., 2013), a specific GR inhibitor (Reul et al., 1990), but also by RU28318 (unpublished observations by KH and MI), a specific MR inhibitor (Ulmann et al., 1985), suggesting the involvement of MR as well as GR. In fact, the expression of MR in PC12 cells has been reported (Goto et al., 2009). Whether MR is involved in the regulation of α3 expression in adrenal chromaffin cells or not remains to be explored. One critical difference between adrenal medulla and brain may be the existence of a higher glucocorticoid level in the extracellular space of adrenal medulla (Wurtman, 2002). Thus, based on the fingdings in PC12 cells, glucocorticoids likely preserve α3 expression in adrenal chromaffin cells. Recently, a decrease in intracellular Cl− concentration ([Cl−]i) has been shown to be involved in the substitution of α1 for α3 (Succol et al., 2012). The value of [Cl−]i in neurons is determined by a balance between Na+, K+, Cl−-cotransporter type 1 (NKCC1) and K+, Cl−-cotransporter type 2 (KCC2), which are responsible for import and export of Cl−, respectively (Kaila et al., 2014). Aldosterone has been reported to enhance NKCC1 activity in vascular smooth muscle cells (Ding et al., 2014) with the consequent increase in [Cl−]i (Davis et al., 1993). These results raise the possibility that glucocorticoids enhance NKCC1 activity in chromaffin cells, thereby preserving α3-containing GABAA receptors. A further study will be required to elucidate the molecular mechanism for α3 preservation in chromaffin cells.

Bottom Line: GABA has two actions mediated by GABAA receptors in chromaffin cells: it induces catecholamine secretion by itself and produces an inhibition of synaptically evoked secretion by a shunt effect.This function of GABA may be facilitated by expression of the immature isoforms of GAD and GABAA receptors and the lack of expression of plasma membrane GABA transporters (GATs).In this review, we will consider how the para/autocrine function of GABA is achieved, focusing on the structural and molecular mechanisms for GABA signaling.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Systems Physiology, University of Occupational and Environmental Health School of Medicine Kitakyushu, Japan.

ABSTRACT
Gamma-aminobutyric acid (GABA) is produced not only in the brain, but also in endocrine cells by the two isoforms of glutamic acid decarboxylase (GAD), GAD65 and GAD67. In rat adrenal medullary chromaffin cells only GAD67 is expressed, and GABA is stored in large dense core vesicles (LDCVs), but not synaptic-like microvesicles (SLMVs). The α3β2/3γ2 complex represents the majority of GABAA receptors expressed in rat and guinea pig chromaffin cells, whereas PC12 cells, an immortalized rat chromaffin cell line, express the α1 subunit as well as the α3. The expression of α3, but not α1, in PC12 cells is enhanced by glucocorticoid activity, which may be mediated by both the mineralocorticoid receptor (MR) and the glucocorticoid receptor (GR). GABA has two actions mediated by GABAA receptors in chromaffin cells: it induces catecholamine secretion by itself and produces an inhibition of synaptically evoked secretion by a shunt effect. Allopregnanolone, a neuroactive steroid which is secreted from the adrenal cortex, produces a marked facilitation of GABAA receptor channel activity. Since there are no GABAergic nerve fibers in the adrenal medulla, GABA may function as a para/autocrine factor in the chromaffin cells. This function of GABA may be facilitated by expression of the immature isoforms of GAD and GABAA receptors and the lack of expression of plasma membrane GABA transporters (GATs). In this review, we will consider how the para/autocrine function of GABA is achieved, focusing on the structural and molecular mechanisms for GABA signaling.

No MeSH data available.


Related in: MedlinePlus