Intracellular ANG II induces cytosolic Ca2+ mobilization by stimulating intracellular AT1 receptors in proximal tubule cells

Am J Physiol Renal Physiol. 2006 Jun;290(6):F1382-90. doi: 10.1152/ajprenal.00269.2005. Epub 2005 Dec 27.


Intracellular ANG II induces biological effects in nonrenal cells, but it is not known whether it plays a physiological role in renal proximal tubule cells (PTCs). PTCs express angiotensinogen, renin, and angiotensin-converting enzyme mRNAs, suggesting the presence of high levels of intracellular ANG II. We determined if microinjection of ANG II directly in single PTCs increases intracellular calcium concentration ([Ca2+]i) and, if so, elucidated the cellular mechanisms involved. Changes in [Ca2+]i responses were studied by fluorescence imaging using the Ca2+ indicator fluo 3. ANG II (1 nM) was microinjected directly in the cells, whereas cell-surface angiotensin type 1 (AT1) receptors were blocked by losartan (10 microM). When ANG II (1 nM) was added to the perfusate, there was a marked increase in [Ca2+]i that was blocked by extracellular losartan. With losartan in the perfusate, intracellular microinjection of ANG II elicited a robust increase in cytoplasmic [Ca2+]i that peaked at 30 s (basal: 2.2 +/- 0.3 vs. ANG II: 14.9 +/- 0.4 relative fluorescence units; P < 0.01). Chelation of extracellular Ca2+ with EGTA (2 mM) did not alter microinjected ANG II-induced [Ca2+]i responses (Ca2+ free + ANG II: 12.3 +/- 2.6 relative fluorescence units, not significant vs. ANG II); however, pretreatment with thapsigargin to deplete intracellular Ca2+ stores or with U-73122 to inhibit phospholipase C (1 microM each) markedly attenuated microinjected ANG II-induced [Ca2+]i responses. Combined microinjection of ANG II and losartan abolished [Ca2+]i responses, whereas a combination of ANG II and PD-123319 had no effect. These data demonstrate for the first time that direct microinjection of ANG II in single PTCs increases [Ca2+]i by stimulating intracellular AT1 receptors and releases Ca2+ from intracellular stores, suggesting that intracellular ANG II may play a physiological role in PTC function.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Angiotensin II / analysis
  • Angiotensin II / pharmacology*
  • Animals
  • Blotting, Western
  • Calcium / metabolism*
  • Calcium / pharmacology
  • Cells, Cultured
  • Cytosol / metabolism
  • Enzyme Activation / drug effects
  • Kidney Tubules, Proximal / chemistry
  • Kidney Tubules, Proximal / metabolism*
  • Kidney Tubules, Proximal / ultrastructure
  • Microinjections
  • Rabbits
  • Receptor, Angiotensin, Type 1 / analysis
  • Receptor, Angiotensin, Type 1 / drug effects
  • Receptor, Angiotensin, Type 1 / physiology*
  • Receptor, Angiotensin, Type 2 / drug effects
  • Receptor, Angiotensin, Type 2 / physiology
  • Thapsigargin / pharmacology
  • Type C Phospholipases / metabolism


  • Receptor, Angiotensin, Type 1
  • Receptor, Angiotensin, Type 2
  • Angiotensin II
  • Thapsigargin
  • Type C Phospholipases
  • Calcium