Effects of Salt Loading on Supraoptic Vasopressin Neurons Assessed by ClopHensorN Chloride Imaging.
Authors of this article are:
Balapattabi K, Farmer GE, Knapp BA, Little JT, Bachelor M, Yuan JP, Cunningham JT.
A summary of the article is shown below:
Salt loading (SL) impairs GABAA inhibition of arginine vasopressin (AVP) neurons in the supraoptic nucleus (SON) of hypothalamus. Based on previous studies, we hypothesized that SL activates tyrosine receptor kinase B (TrkB) downregulating the activity of K+ /Cl- co-transporter2 (KCC2) and upregulating Na+ /K+ /Cl- co-transporter1 (NKCC1). These changes in chloride transport would result in increased [Cl- ]i in SON AVP neurons. This study combined virally mediated chloride imaging with ClopHensorN with single cell Western blot analysis (Simple Wes). An adeno associated virus with ClopHensorN and a vasopressin promoter (AAV2-0VP1-ClopHensorN) was bilaterally injected in the SON of adult male Sprague Dawley rats that were either euhydrated (Eu) or salt loaded (SL) for 7 days. Acutely dissociated SON neurons expressing ClopHensorN were tested for decreases or increases in [Cl- ]i in response to focal application of the GABAA agonist muscimol (100uM). SON AVP neurons from Eu rats showed muscimol-induced chloride influx (p<0.05;23/35). SON AVP neurons from SL rats either significantly increased chloride efflux (p<0.05;27/39) or did not change chloride flux (12/39). The SON AVP neurons that responded to muscimol appeared viable and expressed KCC2 and ß-Actin. Neurons that did not respond during chloride imaging did not have KCC2 and ß-Actin protein expression. The KCC2 antagonist (VU0240551,10uM) significantly blocked the chloride influx in cells from Eu rats but did not affect cells from SL rats. A NKCC1 antagonist (Bumetanide,10uM) significantly blocked the chloride efflux in cells from SL rats but had no effect on cells from Eu rats. Blocking NKCC1 using bumetanide had less effect on the muscimol induced Cl- influx in Eu rat neurons compared to the KCC2 antagonist. The TrkB antagonist (AnA-12,50uM) and protein kinase inhibitor (K252a,100nM) each significantly blocked chloride efflux in SON AVP neurons from SL rats. Salt loading increases [Cl- ]i in SON AVP neurons through TrKB-KCC2-NKCC1 dependent mechanism in rats. This article is protected by copyright. All rights reserved.This article is protected by copyright. All rights reserved.
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This article is a good source of information and a good way to become familiar with topics such as: SON and salt loading; chloride imaging; vasopressin.