Background: Early stages of diabetic nephropathy are characterized by alterations of glomerular filtration, increased tubular sodium and water reabsorption, and systemic volume expansion, which may be a major cause for the development of hypertension. As a significant fraction of renal salt and water transport is mediated by the proximal tubular Na<sup>+</sup>/H<sup>+</sup> exchanger NHE3, we investigated its regulation in rats with STZ-induced diabetes mellitus. Methods: Male Sprague-Dawley rats were injected +/– streptozotocin (STZ, 60 mg/kg), and sacrificed after 2, 7 or 14 days. Renal cortical BBM vesicles were prepared to measure Na<sup>+</sup>/H<sup>+</sup> exchange (NHE) activity and NHE3 protein abundance. Cortical NHE3 mRNA was extracted to perform Northern blot analysis. Pharmacological inhibitors were used in vivo and in vitro in order to identify isoform specificity conferring changes in NHE activity mediated by the diabetic milieu. Results: Compared to control rats, STZ rats were clearly hyperglycemic at all time points studied. NHE activity was significantly increased by 40 and 37% in diabetic rats after 7 and 14 days, respectively, but not after 2 days. The increase in Na<sup>+</sup>/H<sup>+</sup> exchange activity was not inhibited by HOE-642 (3 µ M). Administration of exogenous insulin to diabetic rats resulted in lower blood sugars, but not NHE activity. Moreover, serum glucose concentration did not correlate with NHE activity in any subgroup nor in all animals analyzed together. However, in STZ rats supplemented with exogenous insulin NHE activity was positively correlated with serum insulin concentrations (r = 0.86, p < 0.01). In vivo, the increase in NHE activity induced by STZ could be completely inhibited when rats were fed 6 ppm of HOE-642 with the diet over 14 days. The changes in Na<sup>+</sup>/H<sup>+</sup> exchange activity were not paralleled by changes in NHE3 protein or mRNA abundance in diabetic rats at any of the time points investigated. Conclusions: These results suggest that proximal tubular Na/H exchange activity is modified in the early stage of diabetes mellitus.