Regulation of nephron transport, salt-sensitive hypertension and cross-talk between tubules and vessels
We study regulation of salt and water absorption along the nephron, how these processes alter the renal vasculature and how disruptions in this regulation can lead to hypertension, a disease afflicting 25-30% of the U.S. adult population. Many of the hormones and factors that regulate systemic vascular tone also regulate solute and solvent absorption along the nephron including angiotensin II, endothelin and luminal flow. The reactive oxygen species nitric oxide (NO) and superoxide (O2-) are intermediate signaling molecules for many of the final effects of these factors. Thick ascending limbs reabsorb about 30% of the filtered NaCl. NaCl enters the cells via the luminal Na/K/2 Cl cotransporter type 2 (NKCC2) and exits via basolateral Na/K ATPase. Cl exits the cell via basolateral Cl channels and KCl cotransport. Transport defects in this segment are linked to hypo- and hypertension. Currently we are studying various aspects of how NO and O2- regulate salt absorption by thick ascending limbs. Brief descriptions of these projects are below. Click on "Garvin Lab" (above) for more information.
Regulation of thick ascending limb NaCl absorption by nitric oxide (NO) and superoxide (O2-)
Thick ascending limbs reabsorb about 30% of the NaCl filtered by the glomerulus. NO produced by NO synthase 3 (NOS3) inhibits net NaCl reabsorption by reducing NaCl entry into the cell via NKCC2. Luminal flow and endothelin B, angiotensin type 2 and alpha 2 adrenergic receptor agonists stimulate NOS3 activity. In contrast, O2- enhances net NaCl entry by augmenting NKCC2 activity. O2- is produced by NADPH oxidase, an enzyme composed of several subunits that must assemble to form an active enzyme. Cell stretch caused by luminal flow and angiotensin type 1 receptor agonists activate NADPH oxidase. Flow-induced NO inhibits flow-induced assembly of NADPH oxidase, and thus O2- production. Curiously, angiotensin-induced NO does not have this effect.
Thick ascending limb NO production, its effects on NaCl absorption and defects in salt-sensitive hypertension
Shortly after the endothelium-derived relaxing factor nitric oxide (NO) was described, we published the first report showing that NO inhibits Na transport by the renal cortical collecting duct. This seminal paper led to other studies in this nephron segment. Subsequently, defects in the response to or production of NO were found to cause salt-sensitive hypertension and it had been known for some time that thick ascending limb NaCl absorption was elevated in this disease. Consequently, we shifted our attention to thick ascending limbs. We showed that NO produced by NO synthase 3 (NOS3 or eNOS) in this segment acts as an autacoid to inhibit NKCC2 and as a paracrine factor regulating macula densa function, and thus renal vascular resistance. We are now studying the mechanical transduction process whereby luminal flow stimulates NO production, and how salt-sensitive hypertension affects this. Click on "Garvin Lab" at the top of the page for more details.
Inhibition of O2- production by NO
Our studies of NO also led us to investigate the effects of O2- on transport because O2- scavenges NO, and affects ion transporters in neurons. We found that endogenously produced O2- enhanced thick ascending limb NKCC2 activity via activation of protein kinase C (PKC), and elevated luminal flow stimulated O2- production. This led to the question “Does flow-induced O2- simply scavenge all flow-induced NO?” To our surprise we found that flow-induced O2- only scavenges about 25% of flow-stimulated NO. This is in part due to NO and O2- being produced in different compartments and in part due to the fact that NO prevents activation of NADPH oxidase, the source of flow-induced O2-. We are now studying how NO prevents flow-induced O2-. Clcik on "Garvin Lab" at the top of the page for more details.
Regulation of thick ascending limb transport by angiotensin II
Regulation of thick ascending transport by angiotensin II has been controversial with both stimulation and inhibition being reported by the same investigators. We have shown that angiotensin II stimulates O2- via AT1 receptors but also enhances NO via AT2 receptors. We are now studying the net effects of angiotensin II on thick ascending limb NaCl absorption, the roles of NO and O2- and whether they are salt dependent. Click on "Garvin Lab" at the top of the page for more details.
NO transport by aquaporin-1
Our studies of NO and O2- led us to the conclusion that it is unlikely that NO simply diffuses through the lipid bilayer portion of cell membranes as is current dogma. Whereas transport of NO seemed likely for a number of practical and thermodynamic reasons. We have been able to show that NO influx into proteoliposomes containing purified aquaporin-1 (AQP-1) occurs 4 times faster than in liposomes not containing AQP-1. We were also able to show that transport of NO by AQP-1 has physiological significance. NO-dependent relaxation in aortic rings from AQP-1 knockout mice was significantly reduced compared to wild type mice. We are in the process of developing this project.
- Cabral PD, NJ Hong & JL Garvin.ATP mediates flow-induced NO production in thick ascending limbs. Am. J. Physiol. Renal Physiol. 303:F194-200, 2012.
- Hong NJ & JL Garvin.NADPH oxidase 4 mediates flow-induced superoxide production in thick ascending limbs. Am. J. Physiol. Renal Physiol. 303:F1151-6, 2012.
- Cabral PD, NJ Hong & JL Garvin.Shear stress increases nitric oxide production in thick ascending limbs. Am. J. Physiol. Renal Physiol. 299:F1185-92, 2010.
- Herrera M, GB Silva & JL Garvin.Angiotensin II stimulates thick ascending limb superoxide production via protein kinase C(?)-dependent NADPH oxidase activation. J. Biol. Chem. 285:21323-8, 2010.