Novel pH‐dependent Astrocyte‐Neuron Crosstalk in Hippocampal CA1 Region, Not Observed After the Knockout of the Anion Exchanger 3 (AE3)

The anion exchanger 3 (AE3) is highly expressed in CA1 hippocampal neurons but not astrocytes. It acid loads the cytoplasm by facilitating the exchange of intracellular HCO 3 − for extracellular Cl ‐ . In the present study we investigate the role of AE3 activity in astrocyte‐neuron crosstalk in hippocampal CA1 region during metabolic acidosis (MAc). We bathed our cells in a control (Ctrl) solution that has [HCO 3 − ] of 22 mM at 5% CO 2 , which sets pH at 7.4. We used a MAc solution with pH 7.2, lowering [HCO 3 − ] from 22 to 14 mM at a fixed 5% CO 2 . We used ratiometric fluorescence imaging of the pH sensitive dye BCECF to monitor pH i changes during a 7‐min MAc exposure on: [1] a mixed culture of neurons and astrocytes from the CA1 regions from wild‐type (WT) and AE3 knock‐out (AE3 ‐/‐ ) mice, or [2] a culture of astrocytes only from the CA1 regions from WT and AE3 ‐/‐ mice. In the mixed culture, astrocytes' steady‐state pH i is 6.99 ± 0.01 in WT and 6.92 ± 0.01 in AE3 ‐/‐ . The MAc‐induced acidification rate is ‐3.3 × 10 ‐3 ± 9.1 × 10 ‐4 in the WT and ‐1.8 × 10 ‐3 ± 6.7 × 10 ‐4 in the AE3 ‐/‐ . In the astrocytes‐only culture, steady‐state pH i and MAc‐induced acidification rate in the AE3 ‐/‐ remain the same as in the mixed AE3 ‐/‐ culture. However, in the WT astrocytes‐only culture steady‐state pH i (6.91 ± 0.00) and MAc‐induced acidification rate (‐1.7 × 10 ‐3 ± 6.0 × 10 ‐5 ) are similar to the values for AE3 ‐/‐ astrocytes. The higher steady‐state pH i in the WT mixed culture compared to the WT astrocytes‐only culture presumably occurs because neurons send a message to the astrocytes that [1] increases the astrocytes' steady‐state pH i and [2] speeds the MAc‐induced acidification rate. This crosstalk is absent in AE3 ‐/‐ cells, which may explain the well‐known hyper‐excitability of AE3 ‐/‐ neurons.