Osmoregulation Requires Brain Expression of the Renal Na-K-2Cl Cotransporter NKCC2

Konopacka, Agnieszka; Qiu, Jing; Yao, Song T.; Greenwood, Michael; Greenwood, Mingkwan; Lancaster, Thomas; Inoue, Wataru; Mecawi, André Souza; Vechiato, Fernanda Maria Veanholi; Lima, Juliana Bezerra Medeiros de; Coletti, Ricardo; Hoe, See-Ziau; Martin, Andrew M; Lee, Justina; Joseph, Marina; Hindmarch, Charles; Paton, Julian F. R.; Antunes‐Rodrigues, José; Bains, Jaideep S.; Murphy, David

doi:10.1523/jneurosci.4121-14.2015

Cited by 36 publications

(42 citation statements)

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“…NKCC2 is found widespread in intracellular compartments of non-epithelial cells including hypothalamic neurons (Konopacka et al, 2015), insulin-secreting β-cells (Alshahrani et al, 2012; Alshahrani and Di Fulvio, 2012), or COS7 cells in culture (Alshahrani et al, 2015). To identify the intracellular compartments in which NKCC2 is expressed in COS7, triple immunolabeling of NKCC2, integrin β2 (Iβ2), and NKCC1 was performed.…”

Section: Resultsmentioning

confidence: 99%

“…In fact, only ~3% of the total TALH NKCC2 localized to the cell surface under basal conditions (Ortiz, 2006). Moreover, endogenous NKCC2 has been found in the perinuclear region of different cell types such as insulin-secreting β-cells of the endocrine pancreas (Alshahrani et al, 2012) and vasopressinergic/oxytocinergic neurons of the hypothalamus (Konopacka et al, 2015). Interestingly, most of the exogenously transfected NKCC2 localizes to the perinuclear region of immortalized cell lines derived from mouse TALH (mKTAL), opossum kidney (OKP) or HEK293 cells (Benziane et al, 2007; Zaarour et al, 2009, 2011).…”

Section: Discussionmentioning

confidence: 99%

“…Although there is no doubt that NKCC2 is abundantly expressed in the kidney and in cell lines derived from the TALH (Eng et al, 2007) or the macula densa (Fraser et al, 2007), there is growing evidence showing relatively low levels of expression of extra-renal NKCC2. For instance, NKCC2 expression has been reported in enteric neurons (Xue et al, 2009), gastric, intestinal, endolymphatic sac, and olfactory epithelia (Akiyama et al, 2007, 2010; Nickell et al, 2007; Nishimura et al, 2009; Xue et al, 2009; Zhu et al, 2011; Ji et al, 2012), starburst amacrine cells (Gavrikov et al, 2006), chondrocytes (Bush et al, 2010), and endocrine/neuroendocrine cells including insulin secreting β-cells of the pancreas (Corless et al, 2006; Bensellam et al, 2009; Ghanaat-Pour and Sjöholm, 2009; Alshahrani et al, 2012; Alshahrani and Di Fulvio, 2012) and vasopressinergic/oxytocinergic neurons of the supraoptic and paraventricular nuclei (Hindmarch et al, 2006; Konopacka et al, 2015). …”

Section: Introductionmentioning

confidence: 99%

“…In some cell types, NKCC2 is co-expressed with NKCC1, but whether these proteins interact remains to be determined. NKCC2 expression, plasma membrane localization and function all increase in vasopressinergic and oxytocinergic neurons expressing NKCC1 in rats subjected to chronic dehydration (Hindmarch et al, 2006; Konopacka et al, 2015). These data suggest that the Slc12a1 gene is responsive to osmotic stress.…”

Section: Introductionmentioning

confidence: 99%

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Plasma Membrane Targeting of Endogenous NKCC2 in COS7 Cells Bypasses Functional Golgi Cisternae and Complex N-Glycosylation

Kursan

Almiahoub

Almutairi

et al. 2017

Front. Cell Dev. Biol.

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Na+K+2Cl− co-transporters (NKCCs) effect the electroneutral movement of Na+-K+ and 2Cl− ions across the plasma membrane of vertebrate cells. There are two known NKCC isoforms, NKCC1 (Slc12a2) and NKCC2 (Slc12a1). NKCC1 is a ubiquitously expressed transporter involved in cell volume regulation, Cl− homeostasis and epithelial salt secretion, whereas NKCC2 is abundantly expressed in kidney epithelial cells of the thick ascending loop of Henle, where it plays key roles in NaCl reabsorption and electrolyte homeostasis. Although NKCC1 and NKCC2 co-transport the same ions with identical stoichiometry, NKCC1 actively co-transports water whereas NKCC2 does not. There is growing evidence showing that NKCC2 is expressed outside the kidney, but its function in extra-renal tissues remains unknown. The present study shows molecular and functional evidence of endogenous NKCC2 expression in COS7 cells, a widely used mammalian cell model. Endogenous NKCC2 is primarily found in recycling endosomes, Golgi cisternae, Golgi-derived vesicles, and to a lesser extent in the endoplasmic reticulum. Unlike NKCC1, NKCC2 is minimally hybrid/complex N-glycosylated under basal conditions and yet it is trafficked to the plasma membrane region of hyper-osmotically challenged cells through mechanisms that require minimal complex N-glycosylation or functional Golgi cisternae. Control COS7 cells exposed to slightly hyperosmotic (~6.7%) solutions for 16 h were not shrunken, suggesting that either one or both NKCC1 and NKCC2 may participate in cell volume recovery. However, NKCC2 targeted to the plasma membrane region or transient over-expression of NKCC2 failed to rescue NKCC1 in COS7 cells where NKCC1 had been silenced. Further, COS7 cells in which NKCC1, but not NKCC2, was silenced exhibited reduced cell size compared to control cells. Altogether, these results suggest that NKCC2 does not participate in cell volume recovery and therefore, NKCC1 and NKCC2 are functionally different Na+K+2Cl− co-transporters.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Plasma Membrane Targeting of Endogenous NKCC2 in COS7 Cells Bypasses Functional Golgi Cisternae and Complex N-Glycosylation

Kursan

Almiahoub

Almutairi

et al. 2017

Front. Cell Dev. Biol.

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“…GABA is generally inhibitory in mature neurons due to a high expression of the Cl − exporter KCC2. However, in vasopressin-expressing, but not in oxytocin-expressing, magnocellular neurons, GABA is excitatory under different conditions due to a reduced relative contribution of KCC2 compared to the NKCC1 and NKCC2 Cl − importers [44–46]. …”

Section: Electrical Activity Of Vasopressin Neuronsmentioning

confidence: 99%

Regulation of Neuronal Activity in Hypothalamic Vasopressin Neurons

Ohbuchi

Haam

Tasker

2015

IIS

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Vasopressin is a peptide hormone secreted from the posterior pituitary gland in response to various physiological and/or pathological stimuli, including changes in body fluid volume and osmolality and stress exposure. Vasopressin secretion is controlled by the electrical activity of the vasopressinergic magnocellular neurosecretory cells located in the hypothalamic supraoptic nucleus and paraventricular nucleus. Vasopressin release can occur somatodendritically in the hypothalamus or at the level of pituitary axon terminals. The electrical activity of the vasopressin neurons assumes specific patterns of electrical discharge that are under the control of several factors, including the intrinsic properties of the neuronal membrane and synaptic and hormonal inputs. It is increasingly clear that glial cells perform critical signaling functions that contribute to signal transmission in neural circuits. Astrocytes contribute to neuronal signaling by regulating synaptic and extrasynaptic neurotransmission, as well as by mediating bidirectional neuronal-glial transmission. We recently discovered a novel form of neuronal-glial signaling that exploits the full spatial domain of astrocytes to transmit dendritic retrograde signals from vasopressin neurons to distal upstream neuronal targets. This retrograde trans-neuronal-glial transmission allows the vasopressin neurons to regulate their synaptic inputs by controlling upstream presynaptic neuron firing, thus providing a powerful means of controlling hormonal output.

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Neocortical Microdissection at Columnar and Laminar Resolution for Molecular Interrogation

Kole

Celikel

2018

CP Neuroscience

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The heterogeneous organization of the mammalian neocortex poses a challenge for elucidating the molecular mechanisms underlying its physiological processes. Although high‐throughput molecular methods are increasingly deployed in neuroscience, their anatomical specificity is often lacking. In this unit, we introduce a targeted microdissection technique that enables extraction of high‐quality RNA and proteins at high anatomical resolution from acutely prepared brain slices. We exemplify its utility by isolating single cortical columns and laminae from the mouse primary somatosensory (barrel) cortex. Tissues can be isolated from living slices in minutes, and the extracted RNA and protein are of sufficient quantity and quality to be used for RNA sequencing and mass spectrometry. This technique will help to increase the anatomical specificity of molecular studies of the neocortex, and the brain in general, as it is applicable to any brain structure that can be identified using optical landmarks in living slices. © 2018 by John Wiley & Sons, Inc.

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Osmoregulation Requires Brain Expression of the Renal Na-K-2Cl Cotransporter NKCC2

Cited by 36 publications

References 59 publications

Plasma Membrane Targeting of Endogenous NKCC2 in COS7 Cells Bypasses Functional Golgi Cisternae and Complex N-Glycosylation

Plasma Membrane Targeting of Endogenous NKCC2 in COS7 Cells Bypasses Functional Golgi Cisternae and Complex N-Glycosylation

Regulation of Neuronal Activity in Hypothalamic Vasopressin Neurons

Neocortical Microdissection at Columnar and Laminar Resolution for Molecular Interrogation

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