Podocytes possess the complete machinery for glutamatergic signaling, raising the possibility that neuron-like signaling contributes to glomerular function. To test this, we studied mice and cells lacking Rab3A, a small GTPase that regulates glutamate exocytosis. In addition, we blocked the glutamate ionotropic N-methyl-d-aspartate receptor (NMDAR) with specific antagonists. In mice, the absence of Rab3A and blockade of NMDAR both associated with an increased urinary albumin/creatinine ratio. In humans, NMDAR blockade, obtained by addition of ketamine to general anesthesia, also had an albuminuric effect. In vitro, Rab3A-null podocytes displayed a dysregulated release of glutamate with higher rates of spontaneous exocytosis, explained by a reduction in Rab3A effectors resulting in freedom of vesicles from the actin cytoskeleton. In addition, NMDAR antagonism led to profound cytoskeletal remodeling and redistribution of nephrin in cultured podocytes; the addition of the agonist NMDA reversed these changes. In summary, these results suggest that glutamatergic signaling driven by podocytes contributes to the integrity of the glomerular filtration barrier and that derangements in this signaling may lead to proteinuric renal diseases.

A plasma factor displaying permeability activity in vitro and possibly determining proteinuria has been hypothesized in idiopathic focal segmental glomerulosclerosis (FSGS). In vitro permeability activity (P_alb) was determined in sera of five patients with autosomal recessive steroid-resistant nephrotic syndrome (NPHS2), an inherited condition indistinguishable from idiopathic FSGS on clinical grounds, but in which proteinuria is determined by homozygous mutations of podocin, a key component of the glomerular podocyte. All patients had presented intractable proteinuria with nephrotic syndrome; four developed renal failure and received a renal allograft. For comparison, sera from 31 children with nephrotic syndrome were tested. Pretransplant P_alb was high in all cases (mean 0.81 ± 0.06), equivalent to levels observed in idiopathic FSGS. Overall, P_alb did not correlate with proteinuria. The posttransplant outcome was complicated in two patients by recurrence of proteinuria after 10 and 300 d, respectively, that responded to plasmapheresis plus cyclophosphamide. P_alb levels were high at the time of the recurrence episodes and steadily decreased after plasmapheresis, to reach normal levels in the absence of proteinuria after the seventh cycle. In an attempt to explain high P_alb in these patients, putative inhibitors of the permeability activity were studied. Coincubation of serum with homologous nephrotic urine reduced P_alb to 0, whereas normal urine did not determine any change, which suggests loss of inhibitory substances in nephrotic urine. The urinary levels of the serum P_alb inhibitors apo J and apo E were negligible in all cases, thus suggesting that other urinary inhibitors were responsible for the neutralizing effect. These data indicate that P_alb is high in NPHS2, probably resulting from loss of inhibitors in urine. Lack of correlation of P_alb with proteinuria suggests a selective loss of inhibitors. As in idiopathic FSGS, proteinuria may also recur after renal transplantation in NPHS2 patients, and post-transplant proteinuria is associated with high P_alb. The relationship between elevated P_alb and proteinuria in NPHS2 remains to be determined.

OBJECTIVE: To determine whether HIV-1 Tat may directly alter glomerular permeability in HIV-associated nephropathy (HIVAN). DESIGN: Heavy proteinuria is a hallmark of HIVAN. The slit diaphragm is the ultimate glomerular filtration barrier critical for maintaining the efficiency of the ultrafiltration unit of the kidney. In this study, we evaluated the direct effect of Tat protein on the permeability of isolated glomeruli and on the expression of nephrin, the main slit diaphragm component, by human cultured podocytes. METHODS: Permeability was studied by measuring the permeability to albumin in isolated rat glomeruli. We also evaluated the expression of nephrin in human cultured podocytes by using immunofluorescence and Western blot. RESULTS: We found that Tat increased albumin permeability in isolated glomeruli, and rapidly induced the redistribution and loss of nephrin in cultured podocytes. Pretreatment of glomeruli and podocytes with blocking antibodies showed that Tat reduced nephrin expression by engaging vascular endothelial growth factor receptors types 2 and 3 and the integrin alphavbeta3. Pre-incubation of podocytes with two platelet-activating factor (PAF) receptor antagonists prevented the loss and redistribution of nephrin induced by Tat, suggesting that PAF is an intracellular mediator of Tat action. Tat induced a rapid PAF synthesis by podocytes. When podocytes transfected to overexpress PAF-acetylhydrolase, the main catabolic enzyme of PAF, were stimulated with Tat, the redistribution and loss of nephrin was abrogated. CONCLUSION: The present results define a mechanism by which Tat may reduce nephrin expression in podocytes, thus increasing glomerular permeability. This provides new insights in the understanding of HIVAN pathogenesis