Role Of Renal Na⁺/HCO₃^- Co-transporter NBCn1 And NBCn2 In Regulation Of Systemic Acid-base And NaCl Balance

The kidney plays a fundamental role in maintaining the systemic acid-base and salt balance in the body by secreting acids(mainly in the form of NH₄⁺),reabsorbing HCO₃^-and NaCl.Renal dysfunction can cause severe metabolic acidosis,metabolic alkalosis,hypertension and other diseases.The medullary thick ascending limb of the Henle loop(m TAL)is one of central site for regulating NH₄⁺excretion,HCO₃^-and NaCl reabsorption.In the kidney,the Na⁺/HCO₃^–transporters NBCn1(SLC4A7)and NBCn2(SLC4A10)of the SLC4 family have been localized to the basolateral membrane of the m TAL.The physiological roles of NBCn1 and NBCn2 in the kidney are unclear.Human genetics and gene knockout studies have shown that NBCn1 is associated with hypertension.NBCn2 is associated with dysregulation of plasma osmolality and systemic water balance.In this paper,the expression distribution of NBCn1 and NBCn2 in the kidney and the expression of NBCn1 and NBCn2 in renal m TAL by systemic acid-base or electrolyte stimulation were studied to explore the physiological role of NBCn1 and NBCn2 in the regulation of renal acid-base and NaCl reabsorption.Firstly,we identified,by RT-PCR,the splicing variant of NBCn1 in rat kidney.Two variants,NBCn1-G(major one)and NBCn1-I(minor one),were identified in the kidney.Polyclonal antibodies were used to study the distribution of NBCn1 and NBCn2 in renal tissue,and it was found that the expression of NBCn1 and NBCn2 in the external intramedullary band(ISOM)accounted for about 90%of the total expression in the kidney.It is suggested that m TAL is the main expression site of NBCn1 and NBCn2 in the kidney.Secondly,we examined the effects of metabolic acidosis,hypokalemic alkalosis,dexamethasone,Na HCO₃^-induced metabolic alkalosis,and high salt condition on the expression levels of NBCn1 and NBCn2 in the kidney.(1)Metabolic acidosis induced by NH₄Cl.Adding NH₄Cl 7 or 14 days to drinking water and diet induced metabolic acidosis in rats.Under the condition of metabolic acidosis,the kidney can increase NH₄⁺excretion by up regulating ammonia generation.Our western blot indicated that metabolic acidosis resulted in significant up-regulation of protein expression levels of NBCn1 and NBCn2 in renal m TAL.It is suggested that NBCn1 and NBCn2 play an important role in NH₄⁺transport in m TAL.(2)Hypokalemic alkalosis.Hypokalemia upregulates renal ammonia generation and leads to excessive acid secretion,thus causing potassium deficiency alkalosis.We fed rats a low-potassium diet for 7 or 14 days,which resulted in significant up-regulation of the expression levels of NBCn1 and NBCn2 in the kidney.The results also suggest that NBCn1 and NBCn2 may play an important role in NH₄⁺transport in m TAL.(3)Dexamethasone(glucocorticoid analogues).Dexamethasone can cause renal acid-base disorder,induce the upregulation of renal ammonia generation,and lead to the increase of NH₄⁺transport of m TAL.We injected dexamethasone into rats and found that the expressions of NBCn1 and NBCn2 in the renal medulla were significantly up-regulated.The results also support that NBCn1 and NBCn2 may play an important role in NH₄⁺transport in m TAL.(4)Metabolic alkalosis induced by Na HCO₃.Na HCO₃ was added to drinking water and fed to rats for 7 or 14 days to induce metabolic alkalosis.The results showed that the expression levels of NBCn1 and NBCn2 in kidney were significantly up-regulated.The results suggest that NBCn1 and NBCn2 may play an important role in renal HCO₃^-reabsorption.(5)High salt diet.In order to determine whether the up-regulated expression of NBCn1 and NBCn2 caused by Na HCO₃ was the result of excessive Na⁺,we added NaCl to drinking water and fed the rats for 7 or 14 days.The results showed that NaCl also increased the expression of NBCn1 and NBCn2.Cross-comparison showed that the effect of Na HCO₃ was higher than that of NaCl.The results suggest that NBCn1 and NBCn2may play an important role in renal NaCl reabsorption.Based on our results in the present study and observations from previous studies,we propose that:(1)under the conditions of NH₄Cl-induced metabolic acidosis,hypokalemic alkalosis,and treatment with dexamethasone,the uptake of HCO₃^-in basolateral membrane through NBCn1 and NBCn2 was increased to neutralize more H⁺entering the cell,therefore promote the reabsorption of NH₄⁺;(2)NBCn1 and NBCn2 inhibit HCO₃^-reabsorption by opposing the action of basolateral AE2 during Na HCO₃-induced MAlk;and(3)NBCn1 and NBCn2,in concert with AE2,inhibit NaCl reabsorption during Na⁺loads by mediating net NaCl backflux across the basolateral membrane.Therefore,when the acid-base and electrolyte disorder occurs,NBCn1 and NBCn2 will have corresponding changes to participate in the regulation of homeostasis.Our study suggests that hypertension related to NBCn1 and AE2 may have renal pathological mechanism.