Region-specific changes in transient receptor potential vanilloid channel expression in the vasopressin magnocellular system in hepatic cirrhosis-induced hyponatraemia.

The present study aimed to measure the expression of transient receptor potential (TRP) channels in the magnocellular neurones of the paraventricular (PVN) and supraoptic nucleus (SON) in an animal model of hepatic cirrhosis associated with inappropriate vasopressin (AVP) release. In these studies, we used chronic bile duct ligation (BDL) in the rat, which is a commonly used model of hepatic cirrhosis, associated with elevated plasma AVP. The present study tested the hypothesis that changes in TRP vanilloid (TRPV) channel expression may be related to inappropriate AVP release in BDL rats. To test our hypothesis, we utilised laser capture microdissection of AVP neurones in the PVN and SON and western blot analysis from brain punches. Laser capture microdissection and quantitative reverse transcriptase-polymerase chain reaction demonstrated elevated TRPV2 mRNA in the PVN and SON of BDL compared to sham-ligated controls. AVP transcription was also increased as determined using intron specific primers to measure heteronuclear RNA. Immunohistochemistry demonstrated increased AVP and TRPV2 positive cells in both the PVN and SON after BDL. Also, there was an increased co-expression of TRPV2 and AVP cells after BDL. However, there was no change in the colocalisation counts of TRPV2 and oxytocin in both the magnocellular regions evaluated. In the SON but not the PVN, the transcription levels of TRPV4 were also significantly increased in BDL rats. Western blot analysis of punches containing the PVN and SON revealed that TRPV2 protein content was significantly increased in these brain regions in BDL rats compared to sham rats. Our data suggest that regionally specific changes in TRPV expression in the magnocellular neurosecretory cell AVP neurones could alter their osmosensing ability.

Brain-derived neurotrophic factor-tyrosine kinase B pathway mediates NMDA receptor NR2B subunit phosphorylation in the supraoptic nuclei following progressive dehydration.

We studied the effects of water deprivation (WD) on the phosphorylation of tyrosine kinase B (TrkB) and NMDA receptor subunits in the supraoptic nucleus (SON) of the rat. Laser capture microdissection and quantitative reverse transcriptase polymerase chain reaction was used to demonstrate brain-derived neurotrophic factor (BDNF) and TrkB gene expression in vasopressin SON neurones. Immunohistochemistry confirmed BDNF staining in vasopressin neurones, whereas staining for phosphorylated TrkB was increased following WD. Western blot analysis of brain punches containing the SON revealed that tyrosine phosphorylation of TrkB (pTrkBY(515)), serine phosphorylation of NR1 (pNR1S(866) or pNR1) and tyrosine phosphorylation of NR2B subunits (pNR2BY(1472) or pNR2B) were significantly increased in WD animals compared to controls. Access to water for 2 h reduced pTrkBY(515) content to control levels without affecting pNR1 or pNR2B. Four hours of rehydration was needed to reduce pNR1 and pNR2B to control levels. To test whether increased phosphorylation of TrkB in the present study is mediated by BDNF, a group of animals were instrumented with right SON cannula coupled to mini-osmotic pumps filled with vehicle or TrkB-Fc fusion protein, which prevents BDNF binding to TrkB. In the left SON contralateral to the cannula, TrkB phosphorylation was significantly enhanced following WD. Separate analysis of the right SON, which received TrkB-Fc, showed that the TrkB receptor phosphorylation following WD was significantly attenuated. Although increased pNR1S(866) following WD was not affected by local infusion of TrkB-Fc, pNR2BY(1472) was significantly reduced. Co-immunoprecipitation revealed an increased physical interaction between Fyn kinase and NR2B and TrkB in the SON following WD. Thus, activation of TrkB in the SON following WD may affect cellular excitability through the phosphorylation of NR2B subunits.