The astrocytic Na⁺ -HCO₃ ^- cotransporter, NBCe1, is dispensable for respiratory chemosensitivity

The interoceptive homeostatic mechanism that controls breathing, blood gases and acid-base balance in response to changes in CO₂ /H⁺ is exquisitely sensitive, with convergent roles proposed for chemosensory brainstem neurons in the retrotrapezoid nucleus (RTN) and their supporting glial cells. For astrocytes, a central role for NBCe1, a Na⁺ -HCO₃ ^- cotransporter encoded by Slc4a4, has been envisaged in multiple mechanistic models (i.e. underlying enhanced CO₂ -induced local extracellular acidification or purinergic signalling). We tested these NBCe1-centric models by using conditional knockout mice in which Slc4a4 was deleted from astrocytes. In GFAP-Cre;Slc4a4^fl/fl mice we found diminished expression of Slc4a4 in RTN astrocytes by comparison to control littermates, and a concomitant reduction in NBCe1-mediated current. Despite disrupted NBCe1 function in RTN-adjacent astrocytes from these conditional knockout mice, CO₂ -induced activation of RTN neurons or astrocytes in vitro and in vivo, and CO₂ -stimulated breathing, were indistinguishable from NBCe1-intact littermates; hypoxia-stimulated breathing and sighs were likewise unaffected. We obtained a more widespread deletion of NBCe1 in brainstem astrocytes by using tamoxifen-treated Aldh1l1-Cre/ERT2;Slc4a4^fl/fl mice. Again, there was no difference in effects of CO₂ or hypoxia on breathing or on neuron/astrocyte activation in NBCe1-deleted mice. These data indicate that astrocytic NBCe1 is not required for the respiratory responses to these chemoreceptor stimuli in mice, and that any physiologically relevant astrocytic contributions must involve NBCe1-independent mechanisms. KEY POINTS: The electrogenic NBCe1 transporter is proposed to mediate local astrocytic CO₂ /H+ sensing that enables excitatory modulation of nearby retrotrapezoid nucleus (RTN) neurons to support chemosensory control of breathing. We used two different Cre mouse lines for cell-specific and/or temporally regulated deletion of the NBCe1 gene (Slc4a4) in astrocytes to test this hypothesis. In both mouse lines, Slc4a4 was depleted from RTN-associated astrocytes but CO₂ -induced Fos expression (i.e. cell activation) in RTN neurons and local astrocytes was intact. Likewise, respiratory chemoreflexes evoked by changes in CO₂ or O₂ were unaffected by loss of astrocytic Slc4a4. These data do not support the previously proposed role for NBCe1 in respiratory chemosensitivity mediated by astrocytes.