A Cl(-) cotransporter selective for NH(4)(+) over K(+) in glial cells of bee retina.
There appears to be a flux of ammonium (NH(4)(+)/NH(3)) from neurons to glial cells in most nervous tissues. In bee retinal glial cells, NH(4)(+)/NH(3) uptake is at least partly by chloride-dependant transport of the ionic form NH(4)(+). Transmembrane transport of NH(4)(+) has been described previously on transporters on which NH(4)(+) replaces K(+), or, more rarely, Na(+) or H(+), but no transport system in animal cells has been shown to be selective for NH(4)(+) over these other ions. To see if the NH(4)(+)-Cl(-) cotransporter on bee retinal glial cells is selective for NH(4)(+) over K(+) we measured ammonium-induced changes in intracellular pH (pH(i)) in isolated bundles of glial cells using a fluorescent indicator. These changes in pH(i) result from transmembrane fluxes not only of NH(4)(+), but also of NH(3). To estimate transmembrane fluxes of NH(4)(+), it was necessary to measure several parameters. Intracellular pH buffering power was found to be 12 mM. Regulatory mechanisms tended to restore intracellular [H(+)] after its displacement with a time constant of 3 min. Membrane permeability to NH(3) was 13 microm s(-1). A numerical model was used to deduce the NH(4)(+) flux through the transporter that would account for the pH(i) changes induced by a 30-s application of ammonium. This flux saturated with increasing [NH(4)(+)](o); the relation was fitted with a Michaelis-Menten equation with K(m) approximately 7 mM. The inhibition of NH(4)(+) flux by extracellular K(+) appeared to be competitive, with an apparent K(i) of approximately 15 mM. A simple standard model of the transport process satisfactorily described the pH(i) changes caused by various experimental manipulations when the transporter bound NH(4)(+) with greater affinity than K(+). We conclude that this transporter is functionally selective for NH(4)(+) over K(+) and that the transporter molecule probably has a greater affinity for NH(4)(+) than for K(+).
Marcaggi
P
P
Institut National de la Santé et de la Recherche Medicale U394 Neurobiologie intégrative, Institut François Magendie, Bordeaux, France. p.marcaggi@ucl.ac.uk
Coles
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JA
D000136:Acid-Base Equilibrium / Q000502:physiology; D000641:Ammonia / Q000493:pharmacokinetics; D000818:Animals; D001516:Bees; D002021:Buffers; D002352:Carrier Proteins / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007700:Kinetics; D008954:Models, Biological; D009457:Neuroglia / Q000378:metabolism*; D011188:Potassium / Q000493:pharmacokinetics*; D000644:Quaternary Ammonium Compounds / Q000493:pharmacokinetics*; D012160:Retina / Q000166:cytology / Q000378:metabolism*; D027981:Symporters; D000096922:K Cl- Cotransporters
D000641:Ammonia; D002021:Buffers; D002352:Carrier Proteins; D002712:Chlorides; D011188:Potassium; D000644:Quaternary Ammonium Compounds; D027981:Symporters; D000096922:K Cl- Cotransporters