Effects of extracellular metabolic acidosis and out-of-equilibrium CO/HCO solutions on intracellular pH in cultured rat hippocampal neurons.

Metabolic acidosis (MAc)-an extracellular pH (pH) decrease caused by a [HCO ] decrease at constant [CO]-usually causes intracellular pH (pH) to fall. Here we determine the extent to which the pH decrease depends on the pH decrease vs the concomitant [HCO ] decrease. We use rapid-mixing to generate out-of-equilibrium CO/HCO solutions in which we stabilize [CO] and [HCO ] while decreasing pH (pure acidosis, pAc), or stabilize [CO] and pH while decreasing [HCO ] (pure metabolic/down, pMet↓).

Novel RPTPγ and RPTPζ splice variants from mixed neuron-astrocyte hippocampal cultures as well as from the hippocampi of newborn and adult mice.

Receptor protein tyrosine phosphatases γ and ζ (RPTPγ and RPTPζ) are transmembrane signaling proteins with extracellular carbonic anhydrase-like domains that play vital roles in the development and functioning of the central nervous system (CNS) and are implicated in tumor suppression, neurodegeneration, and sensing of extracellular [CO] and [HCO ]. RPTPγ expresses throughout the body, whereas RPTPζ preferentially expresses in the CNS. Here, we investigate differential RPTPγ-RPTPζ expression in three sources derived from a wild-type laboratory strain of C57BL/6 mice: (a) mixed neuron-astrocyte hippocampal (HC) cultures 14 days post isolation from P0-P2 pups; (b) P0-P2 pup hippocampi; and (c) 9- to 12-week-old adult hippocampi.

Dietary sodium enhances the expression of SLC4 family transporters, IRBIT, L-IRBIT, and PP1 in rat kidney: Insights into the molecular mechanism for renal sodium handling.

The kidney plays a central role in maintaining the fluid and electrolyte homeostasis in the body. Bicarbonate transporters NBCn1, NBCn2, and AE2 are expressed at the basolateral membrane of the medullary thick ascending limb (mTAL). In a previous study, NBCn1, NBCn2, and AE2 are proposed to play as a regulatory pathway to decrease NaCl reabsorption in the mTAL under high salt condition.

Potential Novel Role of Membrane-Associated Carbonic Anhydrases in the Kidney.

Carbonic anhydrases (CAs), because they catalyze the interconversion of carbon dioxide (CO) and water into bicarbonate (HCO) and protons (H), thereby influencing pH, are near the core of virtually all physiological processes in the body. In the kidneys, soluble and membrane-associated CAs and their synergy with acid-base transporters play important roles in urinary acid secretion, the largest component of which is the reabsorption of HCO in specific nephron segments. Among these transporters are the Na-coupled HCO transporters (NCBTs) and the Cl-HCO exchangers (AEs)-members of the "solute-linked carrier" 4 (SLC4) family.

Postnatal Exposure to Brief Hypoxia Alters Brain VEGF Expression and Capillary Density in Adult Mice.

Perinatal hypoxia leads to changes in cerebral angiogenesis and persistent structural and functional changes in the adult brain. It may also result in greater vulnerability to subsequent challenges. We investigated the effect of postnatal day 2 (P2) hypoxic preconditioning on adult brain capillary density and brain vascular endothelial growth factor (VEGF) expression in mice.

Distinguishing among HCO 3- , CO 3= , and H + as Substrates of Proteins That Appear To Be "Bicarbonate" Transporters.

Background: Differentiating among HCO 3- , CO 3= , and H + movements across membranes has long seemed impossible. We now seek to discriminate unambiguously among three alternate mechanisms: the inward flux of 2 HCO 3- (mechanism 1), the inward flux of 1 CO 3= (mechanism 2), and the CO 2 /HCO 3- -stimulated outward flux of 2 H + (mechanism 3).

Methods: As a test case, we use electrophysiology and heterologous expression in Xenopus oocytes to examine SLC4 family members that appear to transport "bicarbonate" ("HCO 3- ").

Carbonic anhydrase and soluble adenylate cyclase regulation of cystic fibrosis cellular phenotypes.

Several aspects of the cell biology of cystic fibrosis (CF) epithelial cells are altered including impaired lipid regulation, disrupted intracellular transport, and impaired microtubule regulation. It is unclear how the loss of cystic fibrosis transmembrane conductance regulator (CFTR) function leads to these differences. It is hypothesized that the loss of CFTR function leads to altered regulation of carbonic anhydrase (CA) activity resulting in cellular phenotypic changes.

Carbon dioxide transport across membranes.

Carbon dioxide (CO) movement across cellular membranes is passive and governed by Fick's law of diffusion. Until recently, we believed that gases cross biological membranes exclusively by dissolving in and then diffusing through membrane lipid. However, the observation that some membranes are CO impermeable led to the discovery of a gas molecule moving through a channel; namely, CO diffusion through aquaporin-1 (AQP1).

Carbonic anhydrases enhance activity of endogenous Na-H exchangers and not the electrogenic Na/HCO cotransporter NBCe1-A, expressed in Xenopus oocytes.

Key Points: According to the metabolon hypothesis, direct association of cytosolic carbonic anhydrases (CAs) with the electrogenic Na/HCO cotransporter NBCe1-A speeds transport by regenerating/consuming . The present work addresses published discrepancies as to whether cytosolic CAs stimulate NBCe1-A, heterologously expressed in Xenopus oocytes. We confirm the essential elements of the previous experimental observations, taken as support for the metabolon hypothesis.

Aquaporin-7: A Dynamic Aquaglyceroporin With Greater Water and Glycerol Permeability Than Its Bacterial Homolog GlpF.

oocytes expressing human aquaporin-7 (AQP7) exhibit greater osmotic water permeability and H-glycerol uptake vs. those expressing the bacterial glycerol facilitator GlpF. AQP7-expressing oocytes exposed to increasing extracellular [glycerol] under isosmolal conditions exhibit increasing swelling rates, whereas GlpF-expressing oocytes do not swell at all.

Multiple acid-base and electrolyte disturbances upregulate NBCn1, NBCn2, IRBIT and L-IRBIT in the mTAL.

Key Points: The roles of the Na /HCO cotransporters NBCn1 and NBCn2 as well as their activators IRBIT and L-IRBIT in the regulation of the mTAL transport of NH , HCO , and NaCl are investigated. Dietary challenges of NH Cl, NaHCO or NaCl all increase the abundance of NBCn1 and NBCn2 in the outer medulla. The three challenges generally produce parallel increases in the abundance of IRBIT and L-IRBIT in the outer medulla.

Computational model of electrode-induced microenvironmental effects on pH measurements near a cell membrane.

The mechanism of gas transport across cell membranes remains a topic of considerable interest, particularly regarding the extent to which lipids vs. specific membrane proteins provide conduction pathways. Studies of transmembrane (CO) transport often rely on data collected under controlled conditions, using pH-sensitive microelectrodes at the extracellular surface to record changes due to extracellular CO diffusion and reactions.

Expression, Localization, and Effect of High Salt Intake on Electroneutral Na/HCO Cotransporter NBCn2 in Rat Small Intestine: Implication in Intestinal NaCl Absorption.

The electroneutral Na/HCO cotransporter NBCn2 (SLC4A10) of solute carrier family 4 (SLC4) plays important physiological and pathological roles in the body. Our previous study showed that NBCn2 is expressed on the protein level in the small intestine of rat. Here, by reverse-transcription polymerase chain reaction (PCR), we identified a novel full-length NBCn2 variant, i.

Role of Carbonic Anhydrases and Inhibitors in Acid-Base Physiology: Insights from Mathematical Modeling.

Carbonic anhydrases (CAs) catalyze a reaction fundamental for life: the bidirectional conversion of carbon dioxide (CO) and water (HO) into bicarbonate (HCO) and protons (H). These enzymes impact numerous physiological processes that occur within and across the many compartments in the body. Within compartments, CAs promote rapid H buffering and thus the stability of pH-sensitive processes.

Increased cerebral vascularization and decreased water exchange across the blood-brain barrier in aquaporin-4 knockout mice.

Aquaporin-4 (AQP4) plays an important role in regulating water exchange across the blood-brain barrier (BBB) and brain-cerebrospinal fluid interface. Studies on AQP-4 knockout mice (AQP4-KO) have reported considerable protection from brain edema induced by acute water intoxication and ischemic stroke, identifying AQP4 as a potential target for therapeutic interventions. However, the long-term effects of chronic AQP4 suppression are yet to be elucidated.

Functionalized Phenylbenzamides Inhibit Aquaporin-4 Reducing Cerebral Edema and Improving Outcome in Two Models of CNS Injury.

Cerebral edema in ischemic stroke can lead to increased intracranial pressure, reduced cerebral blood flow and neuronal death. Unfortunately, current therapies for cerebral edema are either ineffective or highly invasive. During the development of cytotoxic and subsequent ionic cerebral edema water enters the brain by moving across an intact blood brain barrier and through aquaporin-4 (AQP4) at astrocyte endfeet.

Linaclotide improves gastrointestinal transit in cystic fibrosis mice by inhibiting sodium/hydrogen exchanger 3.

Gastrointestinal dysfunction in cystic fibrosis (CF) is a prominent source of pain among patients with CF. Linaclotide, a guanylate cyclase C (GCC) receptor agonist, is a US Food and Drug Administration-approved drug prescribed for chronic constipation but has not been widely used in CF, as the cystic fibrosis transmembrane conductance regulator (CFTR) is the main mechanism of action. However, anecdotal clinical evidence suggests that linaclotide may be effective for treating some gastrointestinal symptoms in CF.

Exploring the autoinhibitory domain of the electrogenic Na /HCO transporter NBCe1-B, from residues 28 to 62.

Key Points: Slc4a4 (mouse) encodes at least five variants of the electrogenic sodium/bicarbonate transporter NBCe1. The initial 41 cytosolic amino acids of NBCe1-A and -D are unique; NBCe1-A has high activity. The initial 85 amino acids of NBCe1-B, -C and -E are unique; NBCe1-B and -C have low activity.

A Novel Stopped-Flow Assay for Quantitating Carbonic-Anhydrase Activity and Assessing Red-Blood-Cell Hemolysis.

We report a novel carbonic-anhydrase (CA) assay and its use for quantitating red-blood-cell (RBC) lysis during stopped-flow (SF) experiments. We combine two saline solutions, one containing HEPES/pH 7.03 and the other, ~1% CO/44 mM [Formula: see text]/pH 8.

Na/HCO Cotransporter NBCn2 Mediates HCO Reclamation in the Apical Membrane of Renal Proximal Tubules.

The kidney maintains systemic acid-base balance by reclaiming from the renal tubule lumen virtually all HCO filtered in glomeruli and by secreting additional H to titrate luminal buffers. For proximal tubules, which are responsible for about 80% of this activity, it is believed that HCO reclamation depends solely on H secretion, mediated by the apical Na/H exchanger NHE and the vacuolar proton pump. However, and the proton pump cannot account for all HCO reclamation.

Role of Cl -HCO exchanger AE3 in intracellular pH homeostasis in cultured murine hippocampal neurons, and in crosstalk to adjacent astrocytes.

Key Points: A polymorphism of human AE3 is associated with idiopathic generalized epilepsy. Knockout of AE3 in mice lowers the threshold for triggering epileptic seizures. The explanations for these effects are elusive.

Role of Receptor Protein Tyrosine Phosphatase γ in Sensing Extracellular CO2 and HCO3.

Regulation of blood pH-critical for virtually every facet of life-requires that the renal proximal tubule (PT) adjust its rate of H(+) secretion (nearly the same as the rate of HCO3 (-) reabsorption, JHCO3 ) in response to changes in blood [CO2] and [HCO3 (-)]. Yet CO2/HCO3 (-) sensing mechanisms remain poorly characterized. Because receptor tyrosine kinase inhibitors render JHCO3 in the PT insensitive to changes in CO2 concentration, we hypothesized that the structural features of receptor protein tyrosine phosphatase-γ (RPTPγ) that are consistent with binding of extracellular CO2 or HCO3 (-) facilitate monitoring of blood CO2/HCO3 (-) concentrations.

Reconstitution of CO2 Regulation of SLAC1 Anion Channel and Function of CO2-Permeable PIP2;1 Aquaporin as CARBONIC ANHYDRASE4 Interactor.

Dark respiration causes an increase in leaf CO2 concentration (Ci), and the continuing increases in atmospheric [CO2] further increases Ci. Elevated leaf CO2 concentration causes stomatal pores to close. Here, we demonstrate that high intracellular CO2/HCO3 (-) enhances currents mediated by the Arabidopsis thaliana guard cell S-type anion channel SLAC1 upon coexpression of any one of the Arabidopsis protein kinases OST1, CPK6, or CPK23 in Xenopus laevis oocytes.