Abstract
Binding of [125I] Tyr A14 human insulin ([125I] insulin) was measured at 4°C in glomeruli and pieces of tubule microdissected from collagenase-treated rat kidneys. For glomeruli and all segments tested, total and non specific binding increased linearly with glomeruli number or tubular length. When determined with 4.0 nM labelled hormone, the distribution of specific binding sites (expressed as 10−18 mol [125I] insulin bound per glomerulus or mm tubule length) was as follows: glomerulus, 2.5±0.3; proximal convoluted tubule (PCT), 12.6±0.6; pars recta (PR), 4.0±2.3; thin descending limb (TDL), 0.6±0.2; thin ascending limb (TAL), 0.6±0.2; medullary thick ascending limb (MAL), 0.8±0.1; cortical ascending limb (CAL), 2.1±0.1; distal convoluted tubule (DCT), 5.6±1.1; cortical collecting tubule (CCT), 3.2±0.3 and outer medullary collecting tubule (MCT), 2.3±0.1. Specific [125I] insulin binding to glomeruli and tubule segments was time- and dose-dependent, saturable, reversible after elimination of free labelled ligand, and inhibited by unlabelled human insulin. When analysed in Scatchard and Hill coordinates, the binding data revealed a negative cooperation in the interaction processes between [125I] insulin and glomerular and tubular binding sites, with apparent dissociation constants and Hill coefficients of the following values: glomerulus, 0.6 nM and 0.60; PCT, 10.0 nM and 0.55; MAL, 4.3 nM and 0.80; CAL, 2.0 nM and 0.74; CCT, 7.6 nM and 0.80 and MCT, 1.0 nM and 0.57 respectively. The stereospecificity of nephron binding sites was assessed in competitive experiments showing that unlabelled bovine and procine insulins were as efficient as human insulin for displacing [125I] insulin, whereas A and B chains of insulin and unrelated peptide hormones were almost inactive. These results indicate that the detected [125I] insulin binding sites may correspond to physiological insulin receptors.
Similar content being viewed by others
Abbreviations
- [125I] Insulin:
-
[125I] Tyr A14 human insulin
- PCT:
-
proximal convoluted tubule
- PR:
-
pars recta
- TDL:
-
thin descending limb
- TAL:
-
thin ascending limb
- MAL:
-
medullary thick ascending limb
- CAL:
-
cortical ascending limb
- DCT:
-
distal convoluted tubule
- CCT:
-
cortical collecting tubule
- MCT:
-
outer medullary collecting tubule
References
Baum M (1987) Insulin stimulates volume absorption in the rabbit proximal convoluted tubule. J Clin Invest 79:1104–1109
Bouizar Z, Rostène WH, Moukhtar MS, Milhaud G (1986) Characterization and quantitative topographical distribution of salmon calcitonin-binding sites in rat kidney sections. FEBS Lett 96:19–22
Butlen D, Morel F (1985) Glucagon receptors along the nephron: [125I] glucagon binding in rat tubules. Pflügers Arch 404:348–353
Butlen D, Mistaoui M, Morel F (1987) Atrial natriuretic peptide receptors along the rat and rabbit nephrons: [125I] alpha-rat atrial natriuretic peptide binding in microdissected glomeruli and tubules. Pflügers Arch 408:356–365
Chabardès D, Montégut M, Mistaoui M, Butlen D, Morel F (1987) Atrial natriuretic peptide effects on cGMP and cAMP contents in microdissected glomeruli and segments of the rat and rabbit nephrons. Pflügers Arch 408:366–372
Chobanian MC, Hammerman MR (1987) Insulin stimulates ammoniagenesis in canine renal proximal tubular segments. Am J Physiol 253:F1171-F1177
De Fronzo RA, Cooke CR, Andres R, Faloona GR, Davis PJ (1975) The effect of insulin on renal handling of sodium, potassium, calcium and phosphate in man. J Clin Invest 55:845–855
De Fronzo RA, Goldberg M, Agus ZS (1976) The effects of glucose and insulin on renal electrolyte transport. J Clin Invest 58: 83–90
De Meyts P, Bianco AR, Roth J (1976) Site-site interactions among insulin receptors. Characterization of the negative cooperativity. J Biol Chem 251:1877–1888
De Meyts P, Van Obberghen E, Roth J, Wollmer A, Brandeburg D (1978) Mapping of the residues responsible for the negative cooperativity of the receptor-binding region of insulin. Nature 273:504–509
Denton RM (1986) Early events in insulin actions. In: Greengard P, Robison GA (eds) Advances in cyclic nucleotides and protein phosphorylation research, vol 20. Raven Press, New York, 293
Felder RA, Blecher M, Calcagno PL, Jose PA (1984) Dopamine receptors in the proximal tubule of the rabbit. Am J Physiol 247:F499-F505
Gammeltoft S, Van Obberghen E (1986) Protein kinase activity of the insulin receptor. Biochem J 235:1–11
Goldman J, Carpenter FH (1974) Zinc binding, circular dichroism, and equilibrium sedimentation studies on insulin (bovine) and several of its derivatives. Biochemistry 13:4566–4574
Guntupalli J, Rogers A, Bourke E (1985) Effect of insulin on renal phosphorus handling in the rat: interaction with PTH and nicotinamide. Am J Physiol 249:F610-F618
Hammerman MR (1985) Interaction of insulin with the renal proximal tubular cell. Am J Physiol 249:F1-F11
Hammerman MR, Rogers S, Hansen VA, Gavin JR III (1984) Insulin stimulates Pi transport in brush border vesicles from proximal tubular segments. Am J Physiol 247:E616-E624
Ikeda K, Matsumoto T, Mouta K, Yamato H, Takahashi H, Ezawa I, Ogata E (1987) The role of insulin in the stimulation of renal 1,25-dihydroxyvitamin D synthesis by parathyroid hormone in rats. Endocrinology 121:1721–1726
Imbert M, Chabardès D, Morel F, Montégut M, Clique A (1974) Hormone-sensitive adenylate cyclase in isolated rabbit glomeruli. Mol Cell Endocrinol 1:295–304
Imbert-Teboul M, Chabardès D, Montégut M, Clique A, Morel F (1978) Vasopressin-dependent adenylate cyclase activity in the rat kidney medulla: evidence for two separate sites of action. Endocrinology 102:1254–1261
Jaffa AA, Miller DH, Bailey GS, Chao J, Margolius HS, Mayfield RK (1987) Abnormal regulation of renal kallikrein in experimental diabetes. Effects of insulin on prokallikrein synthesis and activation. J Clin Invest 80:1651–1659
Khadouri C, Barlet-Bas C, Doucet A (1987) Mechanism of increased tubular Na−K-ATPase during streptozotocin-induced diabetes. Pflügers Arch 409:296–301
Kurokawa K, Lerner L (1980) Binding and degradation of insulin by isolated cortical tubules. Endocrinology 106:655–662
Lau K, Guntupalli J, Eby B (1983) Effects of somatostatin on phosphate transport: evidence for the role of basal insulin. Kidney Int 24:10–15
Levitzski A, Koshland DE Jr (1969) Negative cooperativity in enzyme regulations. Proc Natl Acad Sci USA 62:1121–1128
Meezan E, Freychet P (1979) Rat renal glomeruli and tubules have specific insulin receptors of differing affinity. Mol Pharmacol 16:1095–1100
Meezan E, Freychet P (1982) Binding and degradation of [125I]-insulin by renal glomeruli and tubules isolated from rats. Diabetologia 22:276–284
Morel F (1981) Sites of hormone action in the mammalian nephron. Am J Physiol 240:F159-F164
Morel F, Chabardès D, Imbert-Teboul M (1978) Methodology for enzymatic studies of isolated tubular segments: adenylate cyclase. In: Martinez-Maldonado M (ed) Methods in pharmacology, Renal pharmacology, vol 4B. Plenum Press, New York, p 297
Mujais SK, Kauffman S, Katz AI (1986) Angiotensin II binding sites in individual segments of the rat nephron. J Clin Invest 77:315–318
Murayama N, Ruggles BT, Gapstur SM, Werness JL, Dousa TP (1985) Evidence for beta adrenoceptors in proximal tubules. Isoproterenol-sensitive adenylate cyclase in pars recta of canine nephron. J Clin Invest 76:474–481
Nakamura R, Emmanouel DS, Katz AI (1983) Insulin binding sites in various segments of the rabbit nephron. J Clin Invest 72:388–392
Nakamura R, Hayashi M, Emmanouel DS, Katz AI (1986) Sites of insulin and glucagon metabolism in the rabbit nephron. Am J Physiol 250:F144-F150
Northrup TE, Krezowski PA, Palumbo PJ, Kim JK, Hui YSF, Dousa TP (1979) Insulin inhibition of hormone-stimulated protein kinase systems of rat renal cortex. Am J Physiol 236:E649-E654
Pang DT, Shafer JA (1984) Evidence that insulin receptor from human placenta as a high affinity for only one molecule of insulin. J Biol Chem 259:8589–8596
Ross BD, Guder WG (1982) Heterogeneity and compartmentation in the kidney. In: Sies H (ed) Metabolic compartmentation. Academic Press, London. p 363
Tomita K, Pisano JJ (1984) Binding of [3H] bradykinin in isolated nephron segments of the rabbit. Am J Physiol 246:F732-F737
Tremblay J, Gerzer R, Vinay P, Pang SC, Beliveau R, Hamet P (1985) The increase of cGMP by atrial natriuretic factor correlates with the distribution of particulate guanylate cyclase. FEBS Lett 181:17–22
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Butlen, D., Vadrot, S., Roseau, S. et al. Insulin receptors along the rat nephron: [125I] Insulin binding in microdissected glomeruli and tubules. Pflugers Arch. 412, 604–612 (1988). https://doi.org/10.1007/BF00583761
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00583761