Hypertonicity increases urea transport, as well as the phosphorylation and membrane accumulation of UT-A1, the transporter responsible for urea permeability in the inner medullary collect duct (IMCD). Hypertonicity stimulates urea transport through PKC-mediated phosphorylation. To determine whether PKC phosphorylates UT-A1, eight potential PKC phosphorylation sites were individually replaced with alanine and subsequently transfected into LLC-PK1 cells. Of the single mutants, only ablation of the S494 site dampened induction of total UT-A1 phosphorylation by the PKC activator phorbol dibutyrate (PDBu). This result was confirmed using a newly generated antibody that specifically detected phosphorylation of UT-A1 at S494. Hypertonicity increased UT-A1 phosphorylation at S494. In contrast, activators of cAMP pathways (PKA and Epac) did not increase UT-A1 phosphorylation at S494. Activation of both PKC and PKA pathways increased plasma membrane accumulation of UT-A1, although activation of PKC alone did not do so. However, ablating the PKC site S494 decreased UT-A1 abundance in the plasma membrane. This suggests that the cAMP pathway promotes UT-A1 trafficking to the apical membrane where the PKC pathway can phosphorylate the transporter, resulting in increased UT-A1 retention at the apical membrane. In summary, activation of PKC increases the phosphorylation of UT-A1 at a specific residue, S494. Although there is no cross talk with the cAMP-signaling pathway, phosphorylation of S494 through PKC may enhance vasopressin-stimulated urea permeability by retaining UT-A1 in the plasma membrane.

Experimentally derived Raman spectra of three members of the taxane family are presented and discussed. The taxoids investigated were docetaxel (Taxotere  ), baccatin III and cephalotaxine. These analytes were studied because of their potent antitumor capability or they are potential precursors for the synthesis of highly valuable pharmaceutical drugs. Drop coating deposition Raman (DCDR) spectroscopy is utilized in order to examine the vibrational wavenumbers of the powders and solutions, down to concentrations as low as 1 µM. Normal Raman modes of two of the three taxanes are confirmed by comparing experimental spectra with related literature and quantum mechanical ab initio calculations.

Uncontrolled diabetes mellitus results in osmotic diuresis. Diabetic patients have lowered nitric oxide (NO) which may exacerbate polyuria. We examined how lack of NO affects the transporters involved in urine concentration in diabetic animals. Diabetes was induced in rats by streptozotocin. Control and diabetic rats were given L-NAME for 3 weeks. Urine osmolality, urine output, and expression of urea and water transporters and the Na-K-2Cl cotransporter were examined. Predictably, diabetic rats presented with polyuria (increased urine volume and decreased urine osmolality). Although metabolic parameters of control rats were unaffected by L-NAME, treated diabetic rats produced 30% less urine and osmolality was restored. UT-A1 and UT-A3 were significantly increased in diabetic rat inner medulla. While L-NAME treatment alone did not alter UT-A1 or UT-A3 abundance, absence of NO prevented the upregulation of both transporters in diabetic rats. Similarly, AQP2 and NKCC2 abundance was increased in diabetic animals however, expression of these transporters were unchanged by L-NAME treatment of diabetes. Increased expression of the concentrating transporters observed in diabetic rats provides a compensatory mechanism to decrease solute loss despite persistent glycosuria. Our studies found that although diabetic-induced glycosylation remained increased, total protein expression was decreased to control levels in diabetic rats treated with L-NAME. While the role of NO in urine concentration remains unclear, lowered NO associated with diabetes may be deleterious to the transporters’ response to the subsequent osmotic diuresis.

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