ROD PHOTOTRANSDUCTION MODULATED BY BICARBONATE IN THE FROG RETINA - ROLES OF CARBONIC-ANHYDRASE AND BICARBONATE EXCHANGE
ROD PHOTOTRANSDUCTION MODULATED BY BICARBONATE IN THE FROG RETINA - ROLES OF CARBONIC-ANHYDRASE AND BICARBONATE EXCHANGE
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DOI：
10.1113/jphysiol.1990.sp018139
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发表时间：
1990-07-01
影响因子：
5.5
通讯作者：
SHEVCHENKO, T
中科院分区：
文献类型：
--
作者：
DONNER, K;HEMILA, S;SHEVCHENKO, T
1.在青蛙和虎蝾螈的暗适应视网膜中研究了操纵视网膜CO2-HCO 3-和H+通量对视杆光传导的影响。2.从离体感觉视网膜记录视杆细胞对短暂闪光的光反应，作为视网膜电图质量受体电位，并从离体视杆细胞通过吸管技术记录。实验处理为：（1）改变灌注液中的[CO2] + [HCO 3-];（2）应用乙酰唑胺（AAA），其抑制碳酸酐酶（CA）;和（3）应用4，4“-二异硫氰酸基芪-2，2”-二磺酸（DIDS），其阻断跨细胞膜转运HCO 3-的交换机制。3.视杆细胞内部递质环鸟苷酸（cGMP）的浓度是从视网膜的视杆细胞外节层进行生化测定的，视杆细胞外节层已在与电生理学实验中灌注相同的溶液中孵育。4.引入6 mM碳酸氢钠以取代标称无CO2-HCO 3（12 mm磷酸盐或HEPES; [Na+]常数）林格溶液的一半缓冲液，使视杆外段层中的环GMP浓度加倍，并增加了完整视网膜中视杆的饱和响应幅度和相对灵敏度。5.向含碳酸氢盐的林格溶液中引入0·5 mM-AAA加速了饱和响应和灵敏度的增长。孵育的AAA-碳酸氢盐林格氏溶液中的视网膜环GMP的浓度升高9倍相比，磷酸盐控制。6.在吸入吸移管的孤立杆的光电流中未观察到转换为碳酸氢盐-AAA林格溶液的影响，仅外节段突出到灌注液中。AAA的靶点可能是含CA的Muller细胞。7.将DIDS引入灌注液（在正常pH 7 - 5下）引起光响应的连续衰减，其最终完全消除光敏感性。无论是否存在碳酸氢盐和AAA，都会发生衰变。8.当DIDS灌注液的pH升高到8·5时，在DIDS中失去光敏性的杆几乎完全恢复。当用碳酸氢盐林格溶液在恒定pH（7·5）下洗出DIDS时，它们也被回收。9.有人提出，我们所有的治疗最终调节细胞内的pH值的杆，这是由H+泄漏和HCO 3-运输到细胞的相对速率。速率取决于杆周围的这些离子的浓度; AAA可能通过减慢Muller细胞从内层视网膜的酸运输而起作用。细胞内pH值是视杆细胞光响应的幅度和动力学的重要决定因素。
1. Effects on rod phototransduction following manipulation of retinal CO2-HCO3- and H+ fluxes were studied in dark-adapted retinas of the frog and the tiger salamander. 2. Rod photoresponses to brief flashes of light were recorded from the isolated sensory retina as electroretinogram mass receptor potentials and from isolated rods by the suction-pipette technique. The experimental treatments were: (1) varying [CO2] + [HCO3-] in the perfusion fluid; (2) applying acetazolamide (AAA), which inhibits the enzyme carbonic anhydrase (CA); and (3) applying 4,4''-diisothiocyanatostilbene-2,2''-disulphonic acid (DIDS) which blocks exchange mechanisms transporting HCO3- across cell membranes. 3. The concentration of the internal transmitter of the rods, cyclic GMP, was biochemically determined from the rod outer segment layer of retinas that had been incubated in the same solutions as were for perfusion in the electrophysiological experiments. 4. The introduction of 6 mM-sodium bicarbonate to replace half the buffer of a nominally CO2-HCO3--free (12 mm-phosphate or HEPES; [Na+] constant) Ringer solution doubled the cyclic GMP concentration in the rod outer segment layer and increased the saturating response amplitude and the relative sensitivity of rods in the intact retina. 5. The introduction of 0.cntdot.5 mM-AAA into bicarbonate-containing Ringer solution accelerated the growth of saturated responses and sensitivity. Incubation of the retina in AAA-bicarbonate Ringer solution elevated the concentration of cyclic GMP ninefold compared with the phosphate control. 6. No effects of switching to bicarbonate-AAA Ringer solution were observed in the photocurrent of isolated rods drawn into suction pipettes with only the outer segment protruding into the perfusion fluid. The target of AAA is probably the CA-containing Muller cell. 7. The introduction of DIDS into the perfusate (at normal pH 7.cntdot.5) set off a continuous decay of photoresponses which finally abolished light sensitivity completely. The decay proceeded regardless of whether bicarbonate and AAA were present or not. 8. Rods that had lost their photosensitivity in DIDS recovered almost fully when the pH of the DIDS perfusate was raised to 8.cntdot.5. They also recovered when DIDS was washed out with bicarbonate Ringer solution at constant pH (7.cntdot.5). 9. It is proposed that all our treatments ultimately modulate the intracellular pH of the rods which is determined by the relative rates of H+ leakage and HCO3-transport into the cells. The rates depend on the concentrations of these ions around the rods; AAA probably acts by slowing down the acid transport from the inner retina by Muller cells. Intracellular pH is an important determinant of both the amplitude and the kinetics of rod photoresponses.