Kristensen N B, Hansen O, Clausen T
Institute of Physiology, University of Aarhus, Denmark.
Acta Physiol Scand. 1995 Sep;155(1):67-76. doi: 10.1111/j.1748-1716.1995.tb09949.x.
Using the technique of vanadate-facilitated [3H]ouabain binding we have developed a simple and reliable assay for measuring the concentration of [3H]ouabain binding sites in small fresh or frozen biopsies of rumen epithelium papillae. In bovine and ovine rumen epithelium obtained from the cranio-ventral rumen sac the concentration of [3H]ouabain binding sites was 1.6-4.9 nmol g dry wt-1 (n = 32) and 3.7-5.2 nmol g dry wt-1 (n = 6), respectively. When incubated in oxygenated Krebs-Ringer bicarbonate buffer fresh biopsies of rumen epithelium maintained a high K+ and low Na+ content for at least 6 h. Na+ loading of the biopsies induced about 20-fold increase of the Na+, K(+)-pump activity based on measurement of ouabain suppressible net [86Rb+] influx. The ouabain suppressible net influx of [86Rb+] measured in Na+ loaded biopsies showed a close correlation to the [3H]ouabain binding capacity (r = 0.80, P < 0.01) and corresponded to 47 +/- 2% (n = 9) of the theoretical maximum flux rate. The ouabain suppressible net influx of K+ and [86Rb+] were linearly related (r = 0.73; P < 0.001). The net Na+ efflux was 1.21 times the net K+ influx. It is concluded that rumen epithelium has a large capacity for active Na+/K+ transport and that there is agreement between the concentration of [3H]ouabain binding sites in the epithelium and the ouabain suppressible rate of net [86Rb+] influx in Na+ loaded biopsies in spite of some uncertainty about the maximum turnover number of the Na+, K(+)-pump in rumen epithelium.
利用钒酸盐促进的[³H]哇巴因结合技术，我们开发了一种简单可靠的检测方法，用于测量瘤胃上皮乳头新鲜或冷冻小活检组织中[³H]哇巴因结合位点的浓度。在取自颅腹侧瘤胃囊的牛和羊瘤胃上皮中，[³H]哇巴因结合位点的浓度分别为1.6 - 4.9 nmol g干重⁻¹（n = 32）和3.7 - 5.2 nmol g干重⁻¹（n = 6）。当在充氧的 Krebs-Ringer 碳酸氢盐缓冲液中孵育时，瘤胃上皮的新鲜活检组织至少6小时内保持高钾和低钠含量。基于对哇巴因可抑制的净[⁸⁶Rb⁺]内流的测量，活检组织的钠加载诱导钠钾泵活性增加约20倍。在钠加载的活检组织中测量的哇巴因可抑制的[⁸⁶Rb⁺]净内流与[³H]哇巴因结合能力密切相关（r = 0.80，P < 0.01），相当于理论最大通量率的47 ± 2%（n = 9）。哇巴因可抑制的钾和[⁸⁶Rb⁺]净内流呈线性相关（r = 0.73；P < 0.001）。钠的净外流是钾净内流的1.21倍。结论是瘤胃上皮具有大量主动钠/钾转运能力，尽管瘤胃上皮中钠钾泵的最大周转率存在一些不确定性，但上皮中[³H]哇巴因结合位点的浓度与钠加载活检组织中哇巴因可抑制的净[⁸⁶Rb⁺]内流率之间存在一致性。
