John M. Russell

Cloning, characterization, and chromosomal mapping of a human electroneutral Na(+)-driven Cl-HCO3 exchanger.

The electroneutral Na+-driven Cl-HCO3 exchanger is a key mechanism for regulating intracellular pH (pH i ) in neurons, glia, and other cells. Here we report the cloning, tissue distribution, chromosomal location, and functional characterization of the cDNA of such a transporter (NDCBE1) from human brain (GenBankTM accession number AF069512). NDCBE1, which encodes 1044 amino acids, is 34% identical to the mammalian anion exchanger (AE2); ∼50% to the electrogenic Na/HCO3 cotransporter (NBCe1) from salamander, rat, and humans; ∼73% to mammalian electroneutral Na/HCO3 cotransporters (NBCn1); 71% to mouse NCBE; and 47% to a Na+-driven anion exchanger (NDAE1) fromDrosophila. Northern blot analysis of NDCBE1 shows a robust ∼12-kilobase signal in all major regions of human brain and in testis, and weaker signals in kidney and ovary. This human gene (SLC4A8) maps to chromosome 12q13. When expressed inXenopus oocytes and running in the forward direction, NDCBE1 is electroneutral and mediates increases in both pH i and [Na+] i (monitored with microelectrodes) that require HCO 3 − and are blocked by 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS). The pH i increase also requires extracellular Na+. The Na+:HCO 3 − stoichiometry is 1:2. Forward-running NDCBE1 mediates a36Cl efflux that requires extracellular Na+ and HCO 3 − and is blocked by DIDS. Running in reverse, NDCBE1 requires extracellular Cl−. Thus, NDCBE1 encodes a human, electroneutral Na+-driven Cl-HCO3 exchanger.

Activation of Na+,K+,Cl- cotransport in squid giant axon by extracellular ions: evidence for ordered binding.

Activation of the influx mode of the Na+,K+,Cl- cotransporter (NKCC) by extracellular Na+, K+ and Cl- was studied using the internally dialyzed squid giant axon. Cooperative interactions among the three transported ions were assessed using ion activation of NKCC-mediated 36Cl influx under two sets of experimental conditions. The first, or control condition, used high, non-limiting concentrations of two of the cotransported ions (the co-ions) while activating cotransport with the third ion. Under this non-limiting co-ion condition the calculated Vmax of the cotransporter was between 57 and 60 pmol/cm2/s. The apparent activation (KApp, or half-saturation) constants were: K+, 9 mM; Na+, 52 mM; and Cl-, 146 mM. The second condition used limiting co-ion concentration conditions. In this case, activation by each ion was determined when one of the other two co-ions was present at or near its apparent half-saturation concentration as determined above. Under these limiting conditions, the KApp values for all three co-ions were significantly increased regardless of which co-ion was present at a limiting concentration. The effects on the apparent Vmax were more complicated. When K+ was the limiting co-ion, there was little effect on the Vmax for Na+ or Cl- activation. In contrast, limiting concentrations of Na+ or Cl- both resulted in a large reduction of the apparent Vmax when activating with the other two co-ions. These results are consistent with an ordered binding mechanism for the NKCC in which K+ binds before Na+ or Cl-. Physiological implications for these results are discussed.

Sodium-Coupled Chloride Cotransporters: Discovery and Newly Emerging Concepts

This chapter examines that sodium-coupled chloride cotransporters (NCCCs) are secondary active transporters. Secondary active transport is a remarkable and widespread cellular mechanism used to move solutes across biological membranes against their concentration and/or electrochemical gradients using the potential energy stored in the gradient of another solute, usually that of sodium. The chapter also describes the function of the Na+-coupled cotransporters. At the most fundamental level, both the NKCCs and the NCC act to move Na + and Clˉ across the plasmalemma into cells. Although the NCC was one of the first of this family to undergo functional characterization, detailed functional studies on this transporter have proven difficult owing to the often complicated anatomy of its locations.

The Sodium-Potassium-Chloride Cotransporter, Human Cytomegalovirus and the Cell Cycle

This chapter defines the basics and pathology of human cytomegalovirus (HCMV) infection. HCMV infection is widespread, affecting 50–90% of the adult population. It has the potential to be deadly under certain conditions. The most obvious morphological characteristic of infection with this pathogen is cell swelling, termed cytomegaly, a 2–3-fold enlargement of the host cell. It is this cell enlargement feature that led to the original interest in the possible role of NKCC in the HCMV infection cycle. The virus is spread by contact with body fluids such as blood, saliva, semen, tears, breast milk and vaginal secretions. HCMV, a beta-herpes virus, is an opportunistic virus like other members of the Herpes family. Following the primary infection, it remains latent, hidden in cells of the salivary glands, kidneys, bone cells as well as in blood cells such as lymphocytes and macrophages.

Sodium-Coupled Chloride Cotransporters

This chapter examines that sodium-coupled chloride cotransporters (NCCCs) are secondary active transporters. Secondary active transport is a remarkable and widespread cellular mechanism used to move solutes across biological membranes against their concentration and/or electrochemical gradients using the potential energy stored in the gradient of another solute, usually that of sodium. The chapter also describes the function of the Na+-coupled cotransporters. At the most fundamental level, both the NKCCs and the NCC act to move Na + and Clˉ across the plasmalemma into cells. Although the NCC was one of the first of this family to undergo functional characterization, detailed functional studies on this transporter have proven difficult owing to the often complicated anatomy of its locations.