Soluble adenylyl cyclase is an acid-base sensor in rainbow trout red blood cells that regulates intracellular pH and haemoglobin–oxygen binding

Aim

To identify the physiological role of the acid-base sensing enzyme, soluble adenylyl cyclase (sAC), in red blood cells (RBC) of the model teleost fish, rainbow trout.

Methods

We used: (i) super-resolution microscopy to determine the subcellular location of sAC protein; (ii) live-cell imaging of RBC intracellular pH (pH_i) with specific sAC inhibition (KH7 or LRE1) to determine its role in cellular acid-base regulation; (iii) spectrophotometric measurements of haemoglobin–oxygen (Hb-O₂) binding in steady-state conditions; and (iv) during simulated arterial-venous transit, to determine the role of sAC in systemic O₂ transport.

Results

Distinct pools of sAC protein were detected in the RBC cytoplasm, at the plasma membrane and within the nucleus. Inhibition of sAC decreased the setpoint for RBC pH_i regulation by ~0.25 pH units compared to controls, and slowed the rates of RBC pH_i recovery after an acid-base disturbance. RBC pH_i recovery was entirely through the anion exchanger (AE) that was in part regulated by HCO₃⁻-dependent sAC signaling. Inhibition of sAC decreased Hb-O₂ affinity during a respiratory acidosis compared to controls and reduced the cooperativity of O₂ binding. During in vitro simulations of arterial-venous transit, sAC inhibition decreased the amount of O₂ that is unloaded by ~11%.

Conclusion

sAC represents a novel acid-base sensor in the RBCs of rainbow trout, where it participates in the modulation of RBC pH_i and blood O₂ transport though the regulation of AE activity. If substantiated in other species, these findings may have broad implications for our understanding of cardiovascular physiology in vertebrates.