Effects of noradrenaline on intracellular pH in acutely dissociated adult rat hippocampal CA1 neurones

1 We examined the effects of noradrenaline on steady‐state intracellular pH (pHi) and the recovery of pHi from internal acid loads imposed by the NH4+ prepulse technique in hippocampal CA1 neurones acutely dissociated from adult rats. 2 Under nominally HCO3−‐free conditions, acid extrusion was accomplished by a Na+‐dependent mechanism, probably the amiloride‐insensitive variant of the Na+‐H+ exchanger previously characterized in both fetal and adult rat hippocampal neurones. In the presence of external HCO3−, acid extrusion appeared to be supplemented by a Na+‐dependent HCO3−‐Cl− exchanger, the activity of which was dependent upon the absolute level of pHi. 3 Noradrenaline evoked a concentration‐dependent and sustained rise in steady‐state pHi and increased rates of pHi recovery from imposed intracellular acid loads. The effects of noradrenaline were not dependent upon the presence of external HCO3− but were blocked by substituting external Na+ with N‐methyl‐D‐glucamine, suggesting that noradrenaline acts to increase steady‐state pHi by increasing the activity of the Na+‐H+ exchanger. 4 The effects of noradrenaline on steady‐state pHi and on rates of pHi recovery from imposed acid loads were mimicked by β1‐ and β2‐, but not α‐, adrenoceptor agonists. The β‐adrenoceptor antagonist propranolol blocked the ability of noradrenaline to increase both steady‐state pHi and rates of pHi recovery from acid loads. 5 The effects of noradrenaline on steady‐state pHi and on pHi recovery rates following acid loads were not dependent on changes in [Ca2+]i. However, the effects of noradrenaline were blocked by pre‐treatment with the adenylate cyclase inhibitor 2′,5′‐dideoxyadenosine and the cAMP‐dependent protein kinase inhibitors Rp‐adenosine‐3′,5′‐cyclic monophosphorothioate (sodium salt; Rp‐cAMPS) and N‐[2‐(p‐bromocinnamylamino)ethyl]‐5‐isoquinolinesulphonamide (H‐89). 6 Forskolin, an activator of endogenous adenylate cyclase, and 3‐isobutyl‐1‐methylxanthine, a phosphodiesterase inhibitor, mimicked the ability of noradrenaline to increase both steady‐state pHi and rates of pHi recovery from imposed acid loads, as did Sp‐cAMPS, a selective activator of cAMP‐dependent protein kinase. The effect of forskolin on steady‐state pHi was blocked by pre‐treatment with Rp‐cAMPS whereas the effect of Sp‐cAMPS was enhanced by pre‐treatment with the protein phosphatase inhibitor, okadaic acid. 7 Noradrenaline also increased steady‐state pHi and rates of pHi recovery from imposed acid loads in cultured postnatal rat hippocampal neurones. In this preparation, the effects of noradrenaline were occluded by 18–24 h pre‐treatment with cholera toxin. 8 We conclude that noradrenaline increases the activity of the Na+‐H+ exchanger in rat hippocampal neurones, probably by inducing an alkaline shift in the pHi dependence of the antiport, thereby raising steady‐state pHi. The effects of noradrenaline are mediated by β‐adrenoceptors via a pathway which involves the α‐subunit of the stimulatory G‐protein Gs (Gsα), adenylate cyclase, cAMP and the subsequent activation of cAMP‐dependent protein kinase which, in turn, may phosphorylate the exchange mechanism.