title: Diabetes-induced activation of protein kinase C inhibits store-operated Ca2+ uptake in rat retinal microvascular smooth muscle.
abstract: OBJECTIVE
  To assess the effects of diabetes-induced activation of protein kinase C (PKC) on voltage-dependent and voltage-independent Ca2+ influx pathways in retinal microvascular smooth muscle cells.
  
  
  METHODS
  Cytosolic Ca2+ was estimated in freshly isolated rat retinal arterioles from streptozotocin-induced diabetic and non-diabetic rats using fura-2 microfluorimetry. Voltage-dependent Ca2+ influx was tested by measuring rises in [Ca2+]i with KCl (100 mmol/l) and store-operated Ca2+ influx was assessed by depleting [Ca2+]i stores with Ca2+ free medium containing 5 micromol/l cyclopiazonic acid over 10 min and subsequently measuring the rate of rise in Ca2+ on adding 2 mmol/l or 10 mmol/l Ca2+ solution.
  
  
  RESULTS
  Ca2+ entry through voltage-dependent L-type Ca2+ channels was unaffected by diabetes. In contrast, store-operated Ca2+ influx was attenuated. In microvessels from non-diabetic rats 20 mmol/l D-mannitol had no effect on store-operated Ca2+ influx. Diabetic rats injected daily with insulin had store-operated Ca2+ influx rates similar to non-diabetic control rats. The reduced Ca2+ entry in diabetic microvessels was reversed by 2-h exposure to 100 nmol/l staurosporine, a non-specific PKC antagonist and was mimicked in microvessels from non-diabetic rats by 10-min exposure to the PKC activator phorbol myristate acetate (100 nmol/l). The specific PKCbeta antagonist LY379196 (100 nmol/l) also reversed the poor Ca2+ influx although its action was less efficacious than staurosporine.
  
  
  CONCLUSIONS
  These results show that store-operated Ca2+ influx is inhibited in retinal arterioles from rats having sustained increased blood glucose and that PKCbeta seems to play a role in mediating this effect.
authors.affiliation: Department of Ophthalmology and Vision Sciences, The Queen's University of Belfast, Institute of Clinical Sciences, The Royal Victoria Hospital, Grosvenor Road, Belfast, N. Ireland, UK.
authors.forename: T M
authors.initials: TM
authors.lastname: Curtis
authors.forename: E H
authors.initials: EH
authors.lastname: Major
authors.forename: E R
authors.initials: ER
authors.lastname: Trimble
authors.forename: C N
authors.initials: CN
authors.lastname: Scholfield
mesh_terms: D000818:Animals; D001160:Arterioles / Q000378:metabolism; D001692:Biological Transport; D001786:Blood Glucose / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology* / Q000473:pathology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D008353:Mannitol / Q000494:pharmacology; D008833:Microcirculation / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology / Q000648:ultrastructure; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D009183:Mycotoxins / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012016:Reference Values; D012171:Retinal Vessels / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology / Q000648:ultrastructure; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D001786:Blood Glucose; D004791:Enzyme Inhibitors; D007211:Indoles; D009183:Mycotoxins; D008353:Mannitol; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; C000543:cyclopiazonic acid