acase@tulane.edu

Hydronephrosis can be defined most easily as an accumulation of fluid in the renal pelvicalyceal system and often is accompanied by a significant reduction in glomerular filtration rate (GFR). In rats with congenital unilateral (right sided) hydronephrosis, treatment with either the angiotensin II receptor antagonist saralasin or the thromboxane A$\sb2$ receptor antagonist SQ-29,548 returns GFR to normal levels. The current study defines the single nephron hemodynamic defect in congenital hydronephrosis and evaluates the roles of angiotensin II (Ang II) and thromboxane A$\sb2$ (TxA$\sb2)$ in this glomerular derangement. Renal micro- puncture studies were performed on animals with a right hydronephrotic kidney (HYDRO, n = 8), non-affected litter mates (CONTROL, n = 6), and four groups of hydronephrotic animals treated with the following antagonists; the TxA$\sb2$ receptor antagonist SQ-29,548 (SQ, n = 7) either one of two Ang II receptor antagonists, saralasin (SAR, n = 7) or DuP-753 (DUP, n = 7), and combined treatment with DuP-753 and SQ-29,548 (S&D, n = 7). SNGFR was significantly reduced in the hydronephrotic kidney compared to control (17.6 $\pm$ 2.0 vs. 35.9 $\pm$ 3.7 nl/min, respectively). Treatment with SQ-29,548 completely restored single nephron function (29.0 $\pm$ 3.0 nl/min), while saralasin and DuP-753 only partially restored function (25.6 $\pm$ 1.6 and 27.8 $\pm$ 1.4 nl/min respectively). Combined SQ-29,548 and DuP-753 treatment resulted in a full recovery of single nephron function to 32.9 $\pm$ 4.4 nl/min. The glomerular ultrafiltration coefficient (K$\sb{\rm f})$ was reduced approximately 45% when comparing CONTROL (2.84 $\pm$.22 nl/min/mmHg) to HYDRO (1.64 $\pm$.08 nl/min/mmHg). K$\sb{\rm f}$ returned to control in SAR, DUP and SQ, and increased above control in S&D (5.58 $\pm$ 1.6 nl/min/mmHg). There were no differences in $\rm P\sb{GC},\ P\sb{T},\ or\ \bar\pi\sb{GC}$ between any of the groups studied. These data indicate that the reduction in SNGFR in congenital hydronephrosis is precipitated by a fall in K$\sb{\rm f}$ which is mediated by both Ang II and TxA$\sb2.$ The observation of an increase in K$\sb{\rm f}$ above CONTROL with combined SQ-29,548 and DuP-753 treatment suggests that Ang II and TxA$\sb2$ alter the ultrafiltration coefficient via separate mechanisms / acase@tulane.edu

acase@tulane.edu

The properties of voltage- and calcium-gated K+ channels were examined in sympathetic ganglion neurons acutely isolated from Spontaneously Hypertensive Rats (SHR) for alterations consistent with the elevated neuronal excitability associated with the exaggerated sympathetic outflow observed in this model of human essential hypertension Whole-cell voltage clamp recordings revue two contradictory alterations of the A-type K+ (IA) current of SHR neurons. The voltage-dependence of steady state (1.0 s) inactivation was shifted approximately 6 mV to more hyperpolarized potentials. Considered alone, the inactivations shift is consistent with increased neuronal excitability since the availability of IA to inhibit excitability would be decreased. However, the inactivation shift was offset by an increase in current density which was sufficient to normalize the amplitude of IA when compared physiological potentials. The time course of the onset of fast inactivation and recovery from inactivation were not altered, suggesting that the inactivation shift results from changes in the slow, or C-type inactivation mechanism. Preventing the development of hypertension in the SHR by long term angiotensin converting enzyme inhibition initiated at a prehypertensive age did not affect I A inactivation gating, suggesting that the inactivation shift is a primary alteration neuron properties in the SHR. Current density, on the other hand, was normal in adult SHR in which the development of hypertension as prevented, suggesting IA density elevates in the SHR secondarily to elevated arterial pressure Examination of neuronal firing properties under whole-cell current clamp demonstrated that changing the concentration of intracellular Ca2+ by whole-cell dialysis modulates sympathetic neuron excitability. Superior cervical ganglion sympathetic neurons from normotensive rats displayed hyperexcitability after dialysis with a Ca2+-free internal solution, manifested as repetitive spiking during 400 ms injections of depolarizing current. Dialysis with an internal solution containing [Ca2+] i ≈ 100 nM supported the expected phasic firing properties of these normotensive rat paravertebral neurons. However, the hyperexcitability of SHR sympathetic neurons was unexpectedly blunted following dialysis with the Ca2+-containing internal soluton. The nystatin perforated-patch technique was therefore employed to examine cell electrical behavior while preserving the cellular Ca2+ handling mechanisms. In the absence of whole-cell dialysis, a greater percentage of SHR neurons fired repetitively in response to long injections of depolarizing current, compared to neurons front normotensive rats. These data may indicate that the underling cellular mechanism that imparts hyperexcitability to SHR neurons may involve the handling of intracellular Ca2+ ions Given the key roll of large conductance Ca2+-activated K+ (maxi K) channels in the regulation of Ca2+ influx into sympathetic neurons these channels were examined as potential candidates underlying the altered neuronal function in the SHR. Properties of single, native maxi K channels in membrane patches excised from SHR and normotensive rat neurons were found to be similar, suggesting that maxi K channels may not contribute to the hyperexcitability of SHR neurons / acase@tulane.edu

The role of calcium (Ca$\sp{2+})$ in numerous cell functions has been firmly established. Free intracellular Ca$\sp{2+}$ interacts with calcium binding proteins (CaBP's), such as calmodulin (CaM), which serve as intracellular 'calcium receptors' to transmit the Ca$\sp{2+}$ signal. 1,25-dihydroxyvitamin D$\rm\sb3\ \lbrack1,25(OH)\sb2D\rbrack,$ acts through its receptor (VDR) to maintain plasma calcium homeostasis and also increase tissue levels of several vitamin-D regulated CaBP's Previous studies have provided evidence for 1,25(OH)$\sb2$D regulated CaM-binding proteins in the rat kidney cytosol. The main objectives of this dissertation were to establish the principal CaMBP's stimulated by 1,25(OH)$\sb2$D in the rat kidney, identify some of their functional characteristics, and investigate their relationship to other effects of 1,25(OH)$\sb2$D in this system Protracted treatment (7d) of vitamin D-deficient rats with 100ng 1,25(OH)$\sb2$D specifically stimulated four principal CaMBP's (CaMBP-D145, CaMBP-D117, CaMBP-D98, and CaMBP-D77) in the rat kidney cytosol. CaMBP-D98 and CaMBP-D77 were predominantly cytosolic, whilst CaMBP-D117 was present in both cytosolic and microsomal fractions. CaMBP-D145 was present in all the subcellular fractions, but only the cytosolic CaMBP-D145 was 1,25(OH)$\sb2$D-stimulated. Membrane localization studies confirmed the exclusive cytosolic localization of CaMBP-D77 and CaMBP-D98, whilst CaMBP-D117 and CaMBP-D145 were present in both membrane and cytosolic fractions. A vitamin D-independent 145 kDa CaMBP that co-migrated with the CaMBP-D145 was present in the basolateral membrane preparation. CaMBP-D115, CaMBP-D98, and CaMBP-D77 were predominantly localized to the distal tubule, whilst CaMBP-D145 was present in both nephron segments. The time course and dose responsiveness of 1,25(OH)$\sb2$D-stimulation of the different CaMBP-D's was not significantly different from that of 1,25(OH)$\sb2$D-induced upregulation of VDR which is considered a physiological effect of 1,25(OH)$\sb2$D in the kidney. Compared with the other effects of 1,25(OH)$\sb2$D in the rat kidney and plasma, there was a hierarchy of sensitivity to 1,25(OH)$\sb2$D in the following order: CaBP-D28K $>$ plasma calcium $\sim$ renal alkaline phosphatase $>$ VDR upregulation/CaMBP-D stimulation The results are exciting because they suggest that the Ca-CaM signal transduction pathway may, via changes in activity of some acceptor proteins, effect some of the biological effects of 1,25(OH)$\sb2$D. The results are also important in serving as a foundation for further studies into this intriguing field / acase@tulane.edu

The role of citrate in the kidney is two fold: it is a major renal metabolite supplying about 10% of the metabolic needs of the kidney, and citrate is a potent inhibitor of calcium nephrolithiasis. In urine, citrate maintains calcium in a soluble form by complexation thus preventing further growth to stones. The level of urinary citrate is determined by the extent of citrate reabsorption by the apical membrane of the proximal tubule. Citrate reabsorption has been attributed to NaDC-1, the first cloned dicarboxylate transporter from a growing family of dicarboxylate and sulfate transporters. Citrate is mainly trivalent at physiologic pH; but the divalent form is predominantly transported at the apical membrane. Thus citrate uptake at the apical membrane of the proximal tubule is pH sensitive No cell culture models have been available to explore the regulation of citrate transport in a native environment. The present studies utilizing OK cells, provide the first well characterized cell culture model of citrate transport. Many characteristics of citrate transport found in vivo are mimicked by OK cells. For example, citrate uptake increased by subjecting OK cells to acute and chronic acid loads as well as potassium depletion. This effect of acid-base status in OK cells is similar to that found in rats and humans and thus provides a model to study molecular regulation of citrate transport. Although citrate transport in OK cells corresponds to many aspects of citrate transport in vivo, other characteristics suggest that the transporter is not NaDC-1 Unexpectedly, the presence of apical calcium was found to regulate citrate transport, an effect not previously shown at the cellular level. OK cells studied in low calcium solutions demonstrated increased dicarboxylate citrate transport. A further increase in citrate uptake occurred with decreasing levels of magnesium after calcium removal. OK cells grown on permeable supports showed that the effect of calcium in regulating citrate transport occurs predominantly on the apical membrane. Extracellular calcium appears to change the magnitude of citrate transport as well as some intrinsic characteristics In summary these studies suggest that the apical membrane of the proximal tubule expresses a high affinity, calcium sensitive, sodium dependent dicarboxylate cotransporter which regulates urinary citrate excretion / acase@tulane.edu

acase@tulane.edu

The widespread popularity of both sports and caffeine, the caffeine ban in Olympic activities, the lack of understanding of the physiological and psychological effects of caffeine on high-intensity, intermittent exercise, all indicate the need for more definitive information concerning the caffeine exercise relationship. Although it is difficult to assess the metabolic effects of sports under field conditions, similar metabolic and circulatory modifications can be produced by interval exercise on the cycle ergometer (Keul, 1971). The purpose of this study was to determine the influence of caffeine treatment upon the serum free fatty acid and glycerol, blood glucose and lactate, as well as perceived exertion, oxygen cost, oxygen debt, and performance time values beyond those caused by three 30-minute/one-minute alternating work/rest intervals under a no caffeine treatment. / Eight, trained, male volunteers--ages 21 to 33 yrs.--served as subjects for this study. The exercise level for the high-intensity interval protocol was set at 85 to 90 percent of the maximal aerobic capacity determined by performance on a continuous cycle ergometer test. Two ergometer performance tests were administered approximately one week apart. For one test subjects ingested 5 mg/kg of caffeine and for the other a placebo one hour before beginning pedalling at 70 RPM at 85 to 90 percent of their maximum aerobic power. / A double blind counterbalanced design was used. All metabolic and respiratory data were calculated using the Apple IIe Computer (REP-200B Data Acquisition System, Vacumetrics, Inc.). A two-factor Analysis of Variance with repeated measures and paired t tests were used to test all hypotheses (p $<$.05). / Caffeine significantly increased time to exhaustion, serum free fatty acids, post-exercise oxygen cost levels and lower post exercise glucose levels. These findings suggest that the caffeine-induced glycogen sparing appears to enhance performance time during high intensity intermittent exercise. This is evidenced by significantly greater fat mobilization, oxygen cost, and higher blood glucose levels under the caffeine treatment. / Source: Dissertation Abstracts International, Volume: 52-10, Section: B, page: 5200. / Major Professor: Emily M. Haymes. / Thesis (Ph.D.)--The Florida State University, 1991.

Voltage-dependent potassium (VDK) and calcium currents were studied in normal and clonal (GH$\sb3)$ rat pituitary cells that secrete prolactin (termed lactotrophs) using the whole-cell voltage clamp technique. / Normal lactotrophs were found to possess at least three distinct components of VDK current: (1) a low threshold, transient current sensitive to 4-aminopyridine (4-AP) and relatively insensitive to tetraethylammonium (TEA); (2) a high threshold, slowly inactivating current sensitive to both TEA and 4-AP; and (3) a high threshold, relatively sustained current sensitive to TEA but largely resistant to 4-AP. Each component of current was also distinguishable based on deactivation (tail current) kinetics. / GH$\sb3$ cells have at least two components of VDK current: (1) a low threshold, TEA-insensitive, inactivating current, and; (2) a high threshold, TEA-sensitive, sustained current. The inactivating component bears similarities to both the low threshold, transient current (TEA-insensitivity) and the slowly inactivating current (deactivation and recovery from inactivation kinetics) of normal lactotrophs. The sustained current of GH$\sb3$ cells is kinetically and pharmacologically similar to the sustained component of current in normal lactotrophs. / The kinetic and pharmacological properties of the low threshold, transient calcium current of GH$\sb3$ cells were also studied. Two features set it apart from similar currents in other cells: (1) the rate-limiting transition in channel closing is strictly voltage-dependent at least to $-$160 mV, and; (2) the current is quite resistant to nickel, with half-blockade occurring at 770 $\mu$M. / Source: Dissertation Abstracts International, Volume: 53-03, Section: B, page: 1228. / Major Professor: Christopher J. Lingle. / Thesis (Ph.D.)--The Florida State University, 1992.

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