title: Renin-angiotensin system expression and secretory function in cultured human ciliary body non-pigmented epithelium.
abstract: BACKGROUND
  Renin-angiotensin system (RAS) components have been identified in human ciliary body and aqueous humour, pointing to a role for the RAS in the regulation of aqueous humour dynamics. Here, the authors examine the functional expression of a RAS and the effects of angiotensin II (AII) on a signal transduction pathway and ion secretion mechanism in cultured human ciliary body non-pigmented epithelium (HNPE).
  
  
  METHODS
  RAS expression was examined in cultured HNPE cells using polymerase chain reaction (PCR) analysis. Secretory function was determined using spectrofluorescence imaging microscopy to measure cell calcium (Ca(2+)(I)) and volume responses. Single channel patch clamp techniques were employed to investigate ion channel activity.
  
  
  RESULTS
  PCR analysis demonstrated the expression of angiotensinogen and the AT(1b) receptor in HNPE cells. A large conductance potassium (BK) channel (mean 190 (SEM 5.6) pS, n = 22 cells), was observed in plasma membrane patches. This channel was calcium sensitive with channel open probability (Po) increasing with increasing Ca(2+)(I) (K(0.5) 10.79 (0.44) microM Ca(2+), Hill coefficient of 1.04 (0.04)). AII (100 nM) increased the number (N) of active BK channels in HNPE cells and also the probability of channel opening (Po). N.P(o) increased from 0.008 (0.002) to 1.38 (0.4) following the addition of AII (p=0.0064). AII also induced a rapid rise in Ca(2+)(I) from resting values of 112 (14) nM to a peak of 992 (106) nM (p<10(-4)). A simultaneous cell volume reduction of 24.70% (3.34%) (p<10(-4)) occurred during this calcium signal. Losartan (1 microM) significantly blocked the AII induced BK channel activation (p=0.0131), the Ca(2+)(I) response (p<10(-4)), and the AII induced volume effect (p=0.0046).
  
  
  CONCLUSIONS
  It was demonstrated that AII activates a Ca(2+)(I) signalling system which subsequently increases potassium ion channel activity. These effects are accompanied simultaneously by cell volume loss, indicating that AII acts as receptor operated secretagogue in HNPE cells. The ability of an AT(1) receptor antagonist to inhibit these processes may thus offer a new family of pharmaceutical agents to the current armamentarium in the treatment of glaucoma.
authors.affiliation: Wellcome Trust, Cellular Physiology Research Unit, Department of Physiology, National University of Ireland, Cork, Ireland. abcullinane@hotmail.com
authors.forename: A B
authors.initials: AB
authors.lastname: Cullinane
authors.forename: P S
authors.initials: PS
authors.lastname: Leung
authors.forename: J
authors.initials: J
authors.lastname: Ortego
authors.forename: M
authors.initials: M
authors.lastname: Coca-Prados
authors.forename: B J
authors.initials: BJ
authors.lastname: Harvey
mesh_terms: D000804:Angiotensin II / Q000494:pharmacology; D002118:Calcium / Q000502:physiology; D048429:Cell Size / Q000187:drug effects; D002478:Cells, Cultured; D002924:Ciliary Body / Q000166:cytology* / Q000378:metabolism; D004594:Electrophysiology; D006801:Humans; D051036:Large-Conductance Calcium-Activated Potassium Channels; D018408:Patch-Clamp Techniques; D010857:Pigment Epithelium of Eye / Q000166:cytology* / Q000378:metabolism; D024681:Potassium Channels, Calcium-Activated / Q000187:drug effects / Q000502:physiology; D011945:Receptors, Angiotensin / Q000378:metabolism; D012084:Renin-Angiotensin System / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology
chemical_list: D051036:Large-Conductance Calcium-Activated Potassium Channels; D024681:Potassium Channels, Calcium-Activated; D011945:Receptors, Angiotensin; D000804:Angiotensin II; D002118:Calcium