GLOMERULAR MESANGIAL CELL ALTERED CONTRACTILITY IN HIGH GLUCOSE IS CA2+ INDEPENDENT

In diabetes, loss of renal arteriolar smooth muscle cell contractility leads to intraglomerular hypertension. In glomeruli isolated from str eptozotocin (STZ)-induced diabetic rats, the mesangial cells (smooth m uscle-like) display loss of contractile responsiveness to angiotensin II. This study examines the mechanistic relationship between altered m esangial cell contractility and vasopressor hormone-stimulated Ca2+ si gnaling in high glucose. Glomeruli were isolated from normal or STZ-in duced diabetic rats to observe ex vivo mesangial cell contractile func tion. Also, rat mesangial cells were cultured (10-20 passages) in norm al (5.6 mmol/l) or high (10-25.6 mmol/l) glucose for 1-5 days. Reducti on of glomerular volume and decreased planar surface area of cultured mesangial cells in response to vasoconstrictor stimulation over 60 min were measured by videomicroscopy and personal computer-based morphome try. Contraction of glomenruli isolated from STZ-administered rats in response to endothelin (ET)-1 (0.1 mu mol/l) or the Ca2+ ionophore A23 187 (5 mu mol/l) was impaired significantly compared with that in norm al glucose. In the presence of arginine vasopressin (AW) (1.0 mu mol/l ) or ET-1 (0.1 mu mol/l), mesangial cells demonstrated a dose-dependen t loss of contractile response to increasing glucose concentrations (5 .6-25.6 mmol/l) within 24 h of high-glucose exposure, which was sustai ned for 5 days. Mesangial cells in high glucose were consistently smal ler in size compared with those in normal glucose. Mesangial cells wer e preloaded with myo-[2-H-3]inositol and intracellular [H-3] inositol phosphate release in response to AVP (1.0 mu mol/l) was analyzed by Do wex chromatography. Comparing cells in normal (5.6 mmol/l) versus high (25.6 mmol/l) glucose, we observed no significant difference in stimu lated inositol phosphate levels from 10 to 60 s. The Ca2+-signaling re sponse of cultured mesangial cells preloaded with Fura 2 or Indo 1 was measured by spectrofluorometry. After culture in 25.6 mmol/l glucose, 0.1 mu mol/l AVP or 0.1 mu mol/l ET-1 stimulated a cytosolic Ca2+ sig nal, with first and second phases, identical to that exhibited by mesa ngial cells in normal glucose. Fluorescence imaging of cultured mesang ial cell cytoskeletal filamentous actin (F-actin) stained with rhodami ne-phalloidin demonstrated reduced F-actin staining in the basal state and loss of the normal F-actin disassembly response to ET-1 in high g lucose. Glomerular mesangial cells display glucose-induced altered con traction and F-actin disassembly to vasoactive stimuli, which occur in the presence of normal Ca2+ signaling.