Fig. 5

TRPV4-mediated Ca ²⁺ fluctuations activate NKCC1 in a WNK-SPAK-dependent manner. A Efflux of ⁸⁶Rb⁺ from choroid plexus  in control solution, n = 4 (black), in Ca²⁺-free solution, n = 4 (grey), upon TRPV4-activation by GSK with, n = 4 (green) or without Ca²⁺, n = 4 (light green). B GSK-sensitive efflux rate constants for ⁸⁶Rb⁺ with (green) or without (light green) Ca²⁺ present. C Schematic of the hypothesized LPA-induced TRPV4-mediated Ca²⁺ influx regulating WNK-SPAK-mediated phosphorylation of NKKC1. D Transcript abundance of filtered kinases in rat choroid plexus with rank of SPAK and WNK kinases depicted. E Network analysis depicting published protein-protein associations amongst those illustrated in Panel A. F ⁸⁶Rb⁺ efflux from choroid plexus in control settings without, n = 6 (black) or with SPAK inhibition, n = 6 (grey), or upon TRPV4 activation by GSK without, n = 6 (green) or with, n = 6 (light green) SPAK inhibition. Y-axis is the natural logarithm of the amount of left in the choroid plexus at time t (A_t) divided by the amount at time 0 (A₀). G GSK-mediated efflux rate constants for ⁸⁶Rb⁺ without (green) or with SPAK-inhibition (light green). H ⁸⁶Rb⁺ efflux in control settings without, n = 5 (black) or with, n = 5 (grey) WNK inhibition, or upon TRPV4 activation by GSK without, n = 5 (green) or with, n = 5 (light green) WNK inhibition. I GSK-mediated efflux rate constants for ⁸⁶Rb⁺ without (green) or with (light green) WNK-inhibition. J ⁸⁶Rb⁺ efflux in control settings without, n = 5 (black) or with, n = 5 (grey) NKCC1 inhibition by bumetanide (BUM), or upon application of LPA without, n = 5 (orange) or with, n = 5 (black/orange) NKCC1 inhibition (BUM). K BUM-sensitive efflux rate constants for ⁸⁶Rb⁺ in control setting (black) or upon LPA application (orange). Statistical evaluation with Student’s t-test. *P < 0.05; **P < 0.01; ***P < 0.001