In the absence of exogenous glucocorticoids, decreasing media pH (from 7.4 to 6.8) for 24 hours increased the Na+/H+ exchanger 3 (NHE3) activity in opossum kidney (OKP) cells. 10–7 M and 10–8 M hydrocortisone increased NHE3 activity, and in their presence, acid incubation further increased NHE3 activity. Hydrocortisone (10–9 M) had no effect on NHE3 activity, but in its presence, the effect of acid incubation on NHE3 activity increased twofold. Aldosterone (10–8 M) had no effect. In the absence of hydrocortisone, acid incubation increased NHE3 protein abundance by 47%; in the presence of 10–9 M hydrocortisone, acid incubation increased NHE3 protein abundance by 132%. The increase in NHE3 protein abundance was dependent on protein synthesis. However, 10–9 M hydrocortisone did not modify the effect of acid incubation to cause a twofold increase in NHE3 mRNA abundance. In the absence of protein synthesis, 10–9 M hydrocortisone did potentiate an effect of acid on NHE3 activity, which was due to trafficking of NHE3 to the apical membrane. These results suggest that glucocorticoids and acid interact synergistically at the level of NHE3 translation and trafficking.
Increased Na/H antiporter activity has been demonstrated after in vivo chronic metabolic acidosis as well as in vitro acid preincubation of cultured rabbit renal tubule cells. To study the underlying molecular mechanisms of this adaptive increase in Na/H antiporter activity, the present studies examined the effect of low pH media on Na/H antiporter activity and mRNA abundance in cultured renal tubule cells. Na/H antiporter activity was increased by 60% in a mouse renal cortical tubule cell line (MCT), and by 90% in an opossum kidney cell line (OKP) after 24 h of preincubation in acid (low [HCO3]) media. The ethylisopropylamiloride sensitivity of the Na/H antiporters were different in these two cell lines (MCT IC50 = 65 nM; OKP IC50 = 4.5 microM). In MCT cells, Na/H antiporter mRNA abundance measured by RNA blots increased by two- to fivefold after 24 h in low [HCO3] media. Na/H antiporter mRNA abundance was also increased in MCT cells with high CO2 preincubation as well as in rat renal cortex with in vivo chronic acid feeding. In contrast to renal epithelia, acid preincubation of NIH 3T3 fibroblasts led to suppression of Na/H antiporter activity. RNA blots of 3T3 fibroblasts revealed the same size Na/H antiporter transcript as in MCT cells. However, Na/H antiporter mRNA levels were suppressed by acid preincubation. These studies demonstrate differential regulation of Na/H antiporter activity and mRNA abundance in renal epithelial cells and fibroblasts in response to an acidotic environment.
Glucocorticoids play an important role in modulating proximal tubule acidification. Chronic systemic administration of dexamethasone increases the rate of bicarbonate absorption in isolated perfused proximal convoluted tubules and Na+/H+ antiporter activity in renal brush-border membrane vesicles. The proximal tubule expresses mRNA corresponding to two known Na+/H+ antiporter isoforms: NHE-3, an amiloride-resistant apical membrane Na+/H+ antiporter; and NHE-1, an amiloride-sensitive Na+/H+ antiporter found on most mammalian cells. Administration of dexamethasone for 1 and 2 days (600 micrograms/kg twice daily and 2 h before animals were killed) increased NHE-3 mRNA abundance 1.3- and 2.5-fold, respectively, but had no effect on NHE-1 mRNA abundance. Aminoglutethimide-induced glucocorticoid deficiency had no effect on NHE-1 or NHE-3 mRNA abundance. Incubation of proximal tubules for 3 h with 10(-5) M dexamethasone increased proximal tubule Na+/H+ antiporter activity from 0.69 +/- 0.04 to 0.92 +/- 0.03 pH units/min (P < 0.01); however, there was no increase in NHE-3 or NHE-1 mRNA abundance. Similarly, there was no effect on NHE-3 or NHE-1 mRNA abundance in rabbit renal cortex 4 h after intravenous administration of 600 micrograms/kg dexamethasone. Thus chronic dexamethasone increases NHE-3 but not NHE-1 mRNA abundance. The acute increase in Na+/H+ antiporter activity induced by dexamethasone occurs by mechanisms independent of changes in NHE-1 and NHE-3 mRNA abundance.
Rat proximal convoluted tubules were perfused in vivo to examine the active and passive components of chloride absorption. Chloride flux was a linear function of the transepithelial electrochemical driving force, yielding a permeability coefficient of 20.6 X 10(-5) cm/s. In the absence of an electrochemical driving force, chloride absorption persisted at the rate of 131 peq/mm X min, thus demonstrating active absorption of chloride. Addition of luminal cyanide to tubules absorbing chloride inhibited net chloride absorption. In tubules perfused with a low luminal chloride concentration in which there was net chloride secretion, addition of luminal cyanide increased the magnitude of net chloride secretion. These studies demonstrate that transepithelial chloride transport involves two components: a passive paracellular flux and an active transcellular flux. Cyanide affects net chloride flux by inhibiting active transcellular chloride absorption.
Mechanisms for the citraturic response to potassium citrate treatment were sought by assessing renal citrate clearance and acid-base status after oral administration of potassium citrate, potassium bicarbonate, and potassium chloride. After 2 weeks of treatment of eight patients with stones at a dose of 80 meq/day, urinary citrate rose significantly from 2.5 ± 1.6 mmol/day (no drug) to 5.1 ± 1.7 mmol/day with potassium citrate and to 4.5 ± 1.5 mmol/day with potassium bicarbonate (P < 0.05), but did not change significantly with potassium chloride. Citrate clearance increased from 8.0 to 27.4 mL/min with potassium citrate and 25.8 mL/min with potassium bicarbonate (P < 0.05), but did not ir. crease with potassium chloride. Both potassium citrate and potassium bicarbonate significantly raised urinary bicarbonate and decreased urinary ammonium, titratable acid, and net acid excretion. Potassium chloride was without effect. Effects of potassium citrate on urinary citrate, citrate clearance, and acid-base status tended to be more prominent than those of potassium bicarbonate, but these changes were not significant. Thus, the citraturic action of potassium citrate is largely accountable fo r by provision of an alkali load. Potassium itself had no effect in the absence of potassium deficiency.
The NIH has played a pivotal role in shaping biomedical research into the remarkable success that it is today, through both its intramural program and its generous funding of extramural biomedical science. If you think you know the NIH, you will likely still learn much about it from John A. Kastor’s new book, The National Institutes of Health, 1991–2008, which should be required reading for anyone with an interest in this organization that has been so central to biomedical research.
While the book purports to focus on the years 1991–2008, it frequently retreats back in time to critical periods as the NIH evolved into the institution it is today. The book covers the intramural and extramural programs, addresses the complex structure of the NIH, and describes many of the key leaders, here focusing more on recent times. It describes most of the key controversies, though admittedly not with the depth or entertainment value desired by those in search of exciting discord.
Kastor is an accomplished medical historian who has diligently uncovered the facts underlying the history of the NIH. As he notes, his goal is to describe “how the NIH operates, its problems, its finances, its politics, and its structure within the federal government.” The book includes interviews with a myriad of individuals, most of whom have strong, well-informed opinions regarding what is right and wrong with the NIH. The major strength of the book is that it provides a broad and thorough description of the NIH, its history, its organization, and the many critical people who have served there. The issues surrounding the Intramural Research Program are perhaps the most interesting. During the Vietnam War, universities did not have academic programs that could compete with those at the NIH, and a whole generation of outstanding young researchers joined the intramural program for training. Whether they went to the NIH for superior training or to avoid service in the Vietnam War, the result was that the intramural program defined future biomedical science through these investigators and through their legacy, the researchers who subsequently trained in their labs. But is there a need for the intramural program today? Universities now have well-developed research programs, there is no draft, and the intramural program is not attracting the scientists it once did. This is compounded by limits on compensation, severe budget cuts, and conflict-of-interest rules that all contribute to make universities a more welcome place for the biomedical researcher.
The future of the intramural research program is just one of many controversies that are addressed in Kastor’s book. Should there be a graduate school at the intramural program? Should institute directors yield greater power to the director of the NIH? Was Elias Zerhouni’s Roadmap a heroic attempt to insure that the NIH serves its constituency, the American public, or was it just another assault on the R01-supported investigator? Should the NIH fund institute-directed big science, or should it stay focused on supporting the ideas of researchers through investigator-initiated grants? Should the NIH support clinical research, or should it focus on basic research, leaving the former for industry? Are conflict-of-interest rules overly strict or too lenient? Do they undercut the ability of the NIH to attract the best talent, or are they needed to maintain the trust of the public? Then there are the personal scandals that would be expected to occur at any institution as large as the NIH. Kastor covers all of these controversies in an objective fashion, never expressing personal opinions or bias. Though fair, the evenness of his tone frequently leaves the reader wanting something more lurid.
At times the book gets a little dry, especially as it reviews the structure of each of the institutes and centers, but even here there is much useful information, and the descriptions of the institutes with which the reader has personal experience are of interest.
The NIH has undoubtedly been a driving force in shaping the biomedical research industry in the United States. However, budgets have been relatively flat for a number of years, the future is uncertain, and the American biomedical research enterprise is at great risk. How Congress funds biomedical research and how the NIH chooses to manage the funds it receives will determine whether the United States maintains its preeminence in this arena. For those interested in following or influencing these events, The National Institutes of Health, 1991–2008 should be required — and largely enjoyable — reading.
