Activation of TRPV4 channels promotes the loss of cellular ATP in organotypic slices of the mouse neocortex exposed to chemical ischemia

The vertebrate brain has an exceptionally high energy need. During ischemia, intracellular ATP concentrations decline rapidly, resulting in the breakdown of ion gradients and cellular damage. Here, we employed the nanosensor ATeam1.03(YEMK) to analyse the pathways driving the loss of ATP upon transient metabolic inhibition in neurons and astrocytes of the mouse neocortex. We demonstrate that brief chemical ischemia, induced by combined inhibition of glycolysis and oxidative phosphorylation, results in a transient decrease in intracellular ATP. Neurons experienced a larger relative decline and showed less ability to recover from prolonged (>5 min) metabolic inhibition than astrocytes. Blocking voltage-gated Na+ channels or NMDA receptors ameliorated the ATP decline in neurons and astrocytes, while blocking glutamate uptake aggravated the overall reduction in neuronal ATP, confirming the central role of excitatory neuronal activity in the cellular energy loss. Unexpectedly, pharmacological inhibition of transient receptor potential vanilloid 4 (TRPV4) channels significantly reduced the ischemia-induced decline in ATP in both cell types. Imaging with Na+-sensitive indicator dye ING-2 furthermore showed that TRPV4 inhibition also reduced ischemia-induced increases in intracellular Na+. Altogether, our results demonstrate that neurons exhibit a higher vulnerability to brief metabolic inhibition than astrocytes. Moreover, they reveal an unexpected strong contribution of TRPV4 channels to the loss of cellular ATP and suggest that the demonstrated TRPV4-related ATP consumption is most likely a direct consequence of Na+ influx. Activation of TRPV4 channels thus provides a hitherto unacknowledged contribution to the cellular energy loss during energy failure, generating a significant metabolic cost in ischemic conditions.

Automated summary

The vertebrate brain has a high energy requirement, and ischemia can cause a rapid decline in intracellular ATP concentration, leading to cellular damage. In this study, the pathways driving ATP loss during transient metabolic inhibition were analyzed in mouse neocortex neurons and astrocytes using a nanosensor. It was found that brief chemical ischemia resulted in a temporary decrease in intracellular ATP, with neurons being more affected than astrocytes. Inhibition of voltage-gated Na+ channels, NMDA receptors, or transient receptor potential vanilloid 4 (TRPV4) channels reduced the ATP decline, suggesting the involvement of excitatory neuronal activity and TRPV4 channels in cellular energy loss during ischemia.