Gesek F A, White K E
Department of Pharmacology and Toxicology, Dartmouth Medical School, Hanover, New Hampshire 03755-3835, USA.
Am J Physiol. 1997 Jun;272(6 Pt 2):F712-20. doi: 10.1152/ajprenal.1997.272.6.F712.
Renal nerve stimulation or circulating catecholamines activate the beta-adrenergic receptors that mediate direct effects on tubular transport. Three subtypes of beta-adrenergic receptors have been characterized: beta 1, beta 2, and beta 3. beta-Adrenergic-receptor effects on Na+ and Ca2+ transport in distal convoluted tubules (DCT) have not been established. The focus of this study was to 1) identify the subtypes of beta-adrenergic receptors in DCT cells and 2) examine functional responses to beta-receptor activation on adenosine 3',5'-cyclic monophosphate (cAMP) formation and Na+ and Ca2+ entry. To determine the subtypes of beta-receptors present, RNA isolated from immortalized mouse DCT cells was reverse transcribed, and the cDNA was amplified using primers designed to reported sequences for beta 1-, beta 2-, and beta 3-receptor subtypes. Products of the appropriate sizes were obtained with beta 1- and beta 2-primers. No product was observed with primers to the beta 3 sequence. Receptor products were confirmed by sequencing and are identical to reported mouse beta 1- and beta 2-receptor sequence. Receptor binding of[3H]dihydroalprenolol was 123 +/- 13 fmol/mg protein, and a 3:1 ratio of beta 1- to beta 2-receptors was observed with DCT cell membranes. Isoproterenol, a beta-receptor agonist, increased cAMP formation 8.5-fold. Pretreatment with the antagonist propranolol abolished agonist-induced cAMP accumulation. Isoproterenol significantly increased 22Na+ uptake to 345 +/- 23 compared with a basal rate of 256 +/- 12 nmol.min-1.mg protein-1 and was blocked with propranolol and beta 1- and beta 2-selective antagonists. Isoproterenol had no effect on 45Ca2+ entry into DCT cells. In summary, DCT cells express three times more beta 1- than beta 2-receptors and express no detectable beta 3-adrenergic receptors. beta-Receptors couple to adenylyl cyclase, and activation of beta-adrenergic receptors increases Na+ but not Ca2+ entry in DCT cells.
肾神经刺激或循环中的儿茶酚胺会激活β-肾上腺素能受体，这些受体介导对肾小管转运的直接影响。β-肾上腺素能受体已被鉴定出三种亚型：β1、β2和β3。β-肾上腺素能受体对远曲小管（DCT）中Na+和Ca2+转运的影响尚未明确。本研究的重点是：1）鉴定DCT细胞中β-肾上腺素能受体的亚型；2）研究β-受体激活对3',5'-环磷酸腺苷（cAMP）形成以及Na+和Ca2+内流的功能反应。为了确定存在的β-受体亚型，从永生化小鼠DCT细胞中分离的RNA进行逆转录，然后使用针对β1、β2和β3受体亚型的报道序列设计的引物扩增cDNA。用β1和β2引物获得了合适大小的产物。用β3序列的引物未观察到产物。通过测序确认了受体产物，其与报道的小鼠β1和β2受体序列相同。[3H]二氢阿普洛尔的受体结合量为123±13 fmol/mg蛋白，在DCT细胞膜上观察到β1与β2受体的比例为3:1。β-受体激动剂异丙肾上腺素使cAMP形成增加8.5倍。用拮抗剂普萘洛尔预处理可消除激动剂诱导的cAMP积累。与基础速率256±12 nmol·min-1·mg蛋白-1相比，异丙肾上腺素使22Na+摄取显著增加至345±23，并且被普萘洛尔以及β1和β2选择性拮抗剂阻断。异丙肾上腺素对45Ca2+进入DCT细胞没有影响。总之，DCT细胞中β1受体的表达量是β2受体的三倍，且未检测到β3肾上腺素能受体的表达。β-受体与腺苷酸环化酶偶联，β-肾上腺素能受体的激活增加了DCT细胞中的Na+内流，但不增加Ca2+内流。
