Role of Histamine in the Regulation of Calcium and Actin Dynamics in Collecting Duct Cells

Abstract

Background. Histamine is a nitrogen-based molecule that has established involvement in allergic reactions and immune responses. The levels of histamine are increased in renal pathologies, such as nephrotic syndrome, diabetic nephropathy, acute renal failure, and end stage kidney disease. We have previously shown that all 4 histamine receptors (HR1 to HR4) are abundant in the kidney, and histamine exposure can affect levels of intracellular Ca2+ in the collecting ducts. Calcium signaling is an important regulator of actin cytoskeleton dynamics in these cells, and thus, affects their structure and integrity; however, there is a gap in knowledge regarding the role that histamine may play in this mechanism. In this study, we hypothesized that histamine exposure induces Ca2+ release and leads to reorganization of F-actin cytoskeleton in the cortical collecting duct cells. Methods. Immunofluorescent staining of cultured cells, in combination with the Western blotting, were utilized to characterize the renal expression of HRs and enzymes of the histaminergic system (HiS). The effects of histamine were tested in cultured mouse cortical collecting duct cells (mpkCCD). Functional live confocal imaging was used to detect changes in intracellular Ca2+ level (Fluo 8, 490/525 nm). Actin cytoskeleton rearrangements were assessed by F-actin staining with rhodamine-phalloidin. Images were analyzed in FIJI (NIH). Differences between the groups were assessed with 1-way ANOVA; p < 0.05 was considered significant. Results. IHC and Western blotting showed that all four HRs as well as HiS components are expressed in the mpkCCD cells. Acute application of histamine (100 uM) led to an increase in intracellular Ca2+ level which was predominantly regulated by extracellular influx, with a minor component of intracellular store release. Next, we blocked HR1, HR2, HR3, HR4 with loratadine (5uM), ranitidine (10 uM), iodophenpropit (500 nM), and A943931 (100 nM), respectively, to discern the involvement of specific HRs. We demonstrated that HR1, HR3, HR4 but not HR2 are involved in mediating the influx of Ca2+ in response to histamine. When mpkCCD cells were incubated with 100 uM histamine for 4 hours, we observed formation of the F-actin stress-fibers in the submembrane space. Conclusions. We have shown that collecting duct cells express HRs as well as HiS enzymes, indicating presence of a fully functional HiS. Our data revealed that histamine exposure leads to an influx of Ca2+ in these cells, which is mediated via H1R, H3R, and H4R. Additionally, histamine induced actin cytoskeleton rearrangements, suggesting its role in maintenance of renal epithelial cell structure. Our findings highlight the relationship between histamine, calcium regulation, and actin cytoskeleton dynamics in the renal epithelial cells, offering insights into future therapeutic strategies. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.