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J Neurophysiol 84: 281-288, 2000;
0022-3077/00 $5.00
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The Journal of Neurophysiology Vol. 84 No. 1 July 2000, pp. 281-288
Copyright ©2000 by the American Physiological Society

Reversibility and Cation Selectivity of the K+-Clminus Cotransport in Rat Central Neurons

Yasuhiro Kakazu,1,2 Soko Uchida,1 Takashi Nakagawa,2 Norio Akaike,1 and Junichi Nabekura1

 1Cellular and System Physiology and  2Otorhinolaryngology, Graduate School of Medical Science, Kyushu University, Fukuoka 812-8582, Japan

Kakazu, Yasuhiro, Soko Uchida, Takashi Nakagawa, Norio Akaike, and Junichi Nabekura. Reversibility and Cation Selectivity of the K+-Clminus Cotransport in Rat Central Neurons. J. Neurophysiol. 84: 281-288, 2000. The reversibility and cation selectivity of the K+-Cl- cotransporter (KCC), which normally extrudes Cl- out of neurons, was investigated in dissociated lateral superior olive neurons of rats using the gramicidin perforated patch technique. Intracellular Cl- activity (alpha [Cl-]i) was maintained well below electrochemical equilibrium as determined from the extracellular Cl- activity and the holding potential, where the pipette and external solutions contained 150 mM K+ ([K+]pipette) and 5 mM K+ ([K+]o), respectively. Extracellular application of 1 mM furosemide or elevated [K+]o increased alpha [Cl-]i. When the pipette solution contained 150 mM Cs+ ([Cs+]pipette), alpha [Cl-]i increased to a value higher than the passive alpha [Cl-]i. An increase of alpha [Cl-]i with the [Cs+]pipette was not due to the simple blockade of net KCC by the intracellular Cs+ since alpha [Cl-]i, with the pipette solution containing 75 mM Cs+ and 75 mM K+, reached a value between those obtained using the [K+]pipette and the [Cs+]pipette. The higher-than-passive alpha [Cl-]i with the [Cs+]pipette was reduced by 1 mM furosemide, but not by 20 µM bumetanide or Na+-free external solution, indicating that the accumulation of [Cl-]i in the [Cs+]pipette was mediated by a KCC operating in a reversed mode rather than by Na+-dependent, bumetanide-sensitive mechanisms. Replacement of K+ in the pipette solution with either Li+ or Na+ mimicked the effect of Cs+ on alpha [Cl-]i. On the other hand, Rb+ mimicked K+ in the pipette solution. These results indicate that K+ and Rb+, but not Cs+, Li+, or Na+, can act as substrates of KCC in LSO neurons.




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