Abstract Body: Background: Renin secretion from the juxtaglomerular (JG) cells controls circulating Angiotensin II (AngII), a critical blood pressure determinant. AngII is believed to suppress renin release from JG cells to prevent excessive blood pressure elevation. However, a direct negative feedback mechanism has yet to be established in JG cells. In JG cells, Ca²⁺ is a crucial second messenger that governs renin secretion by suppressing cAMP, a potent renin secretion stimulator. Thus, in contrast to most secretory cells, renin secretion from JG cells is inversely related to the intracellular Ca²⁺ level. However, the role of AngII in modulating JG cell Ca²⁺dynamics remains unclear.
Objective: Define intracellular Ca²⁺ responses to AngII and identify the critical channels to suppress renin secretion in JG cells.
Methods: Kidney slices from mice expressing JG-specific GCaMP6f, a genetically encoded Ca²⁺ indicator under the control of the Ren1^c promoter, were imaged ex vivo on a widefield fluorescent microscope for 30 min per experiment (n = 5). In each experiment, slices were continuously perfused with buffer containing variable Ca²⁺ and AngII concentrations, ± specific Ca²⁺ channel inhibitors. Images were captured at 10 Hz and analyzed using Mesmerize Ca²⁺ imaging software to identify and plot fluorescent intensity over time for each JG cell. Additional kidney slices were incubated in 24-well plates with buffer containing vehicle or AngII ± Ca²⁺channel inhibitors for 30 min. ELISA determined renin concentration.
Results: 1) AngII dose-dependently evoked robust and periodic Ca²⁺-oscillations (Ca²⁺ spikes) in JG cells (mean spikes/min; 50 pM: 5.4, 300 pM: 7.7, 3 nM: 13.2, and 300 nM: 17.6). 2) AngII-elicited Ca²⁺ activity was markedly reduced with “Ca²⁺-free” buffer. 3) L-type voltage-dependent Ca²⁺ channel (VDCC) blocker nifedipine moderately decreased AngII-elicited Ca²⁺ spikes, while T-type VDCC blocker TTA-P2 did not. 4) Blocking endoplasmic reticulum (ER) Ca²⁺-release drastically reduced Ca²⁺ activity. 5) AngII dose-dependently decreased renin secretion in kidney slices, congruent with the AngII-elicited Ca²⁺ activity in imaging studies.
Conclusion: AngII dose-dependently and directly elicited large, periodic Ca²⁺-oscillations in JG cells that suppressed renin release. Both extracellular Ca²⁺ influx and intracellular Ca²⁺ release from ER stores are required for sustained Ca²⁺ activity. L-type, but not T-type, Ca²⁺ channels participate in JG cell Ca²⁺-oscillations.