Sympathetic Overactivity In CKD Disrupts Buffering Of Neurotransmission By Endothelium-derived Hyperpolarizing Factor And Enhances Vasoconstriction

BackgroudHypertension commonly complicates chronic kidney disease(CKD).Resistance arteries are vessels with lumen diameters measuring<400 μM when relaxed,and they constitute the major site of generation of vascular resistance.Vascular smooth muscle cells(VSMCs)on the outer side of resistance arteries receive signals from the sympathetic nervous system(SNS)that induce an endothelial cell(EC)anti-contractile response that moderates the vasoconstriction.However,the specific role of this pathway in the enhanced vasoconstriction in CKD is unknown.Here,we test the hypothesis that an impaired anti-contractile mechanism in resistance arteries in CKD enhances sympathetic vasoconstriction.MethodsMice model of hypertensive CKD was conducted by five-sixth nephrectomy(5/6Nx).Neurovascular transmission was assessed in mesenteric resistance arteries(MRAs)from 5/6Nx or sham mice at week 4 in response to activation of SNS by bilateral carotid artery occlusion.Initial contraction and subsequent anti-contractile responses to norepinephrine was determined in isolated MRAs by wire myograph.Expression and function of connexins in myoendothelial junctions(MEJs)were analyzed in a VSMC-EC coculture system.Rescue of Connexin43 expression in MRAs from 5/6Nx mice was performed by transfection of a GV362-Connexin43 plasmid into mice or isolated MRAs.To determine whether the impaired anti-contractile responses to NE in CKD depends on continuous SNS enhancement,the reactivity of isolated MRAs to NE was compared between 5/6Nx mice with and without blocking the SNS outflow by prolonged icv clonidine treatment.ResultsThe CKD(5/6Nx)mice had a progressive increase in MAP from 2-8 weeks,accompanied by a persistent rise in SNS activity and MRA level of norepinephrine(NE)from 2 to 8 weeks.However,the MRA median/lumen ratio was increased significantly only at week 8.This highlights the importance of functional vascular factors in this early phase of hypertension in this model.Neurovascular transmission was assessed in response to carotid artery occlusion in mesenteric artery of 5/6Nx or sham mice at week 4.5/6Nx mice had a great increase in mesenteric vascular resistance and MAP.The reactivity of MRAs to NE was explored to dissociate neurovascular reactivity from hypertension.The tension of MRAs from sham mice rose rapidly("phasic contraction"),followed by a reduction in tension("anti-contractile component")to a steady state("tonic contraction").Although phasic contractions were similar in both groups,the anti-contractile component of neurotransmission in MRAs in 5/6Nx mice was quite curtailed resulting in a greater tonic contraction.This impaired vasodilation during NE application was abolished by blockade of EDH,which indicated the impair of NE-induced EDH response in 5/6Nx MRAs.This impaired anti-contractile response during NE application of MRA in SHAM and 5/6Nx were also abolished by disruption of gap junctions with 18βGA,indicating a critical role of MEJ gap junction in NE-induced EDH response in 5/6Nx MRAs.The expression of Connexin43 was decreased at VSMC and MEJ of 5/6Nx MRA.Restoration of arterial Connexin43 expression in animals with CKD rescued the impaired EDH response to NE,and corrected the enhanced vascular resistance and MAP after carotid artery occlusion.Taken together,these findings imply a critical role of the decreased Connexin43-containing MEJ gap junction in the impaired neurovascular transmission in CKD.The disrupted neurovascular transmission in CKD relies on a continuously enhanced sympathetic discharge.Exposure of VSMCs to norepinephrine for 24 hours in a vascular cell coculture decreased MEJ connexin 43 expression and MEJ gap junction function.Blockade sympathetic discharge could restorate Connexin43 expression at both VSMC and MEJ in CKD,which improved neurovascular transmission.Conclusion5/6Nx mice have enhanced neurovascular transmission and vasoconstriction from an impaired EDH anti-contractile component in advance of vascular structural changes.These neurovascular changes depend on an enhanced SNS discharge that impairs the expression of connexin 43 in gap junctions at the MEJ,thereby interrupting EDH responses that normally moderate vascular tone.Dysregulation of neurovascular transmission may underline the development of hypertension in CKD.