Bradykinin decreases outflow facility in perfused anterior segments and induces shape changes in passaged BTM cells in vitro.
OBJECTIVE
To investigate the effects of bradykinin (BK) on the outflow facility (C) of human and bovine perfused anterior segments, the [Ca2+]i of cultured bovine trabecular meshwork (BTM) cells, and the area and major axis of BTM cells.


METHODS
Cellular studies were performed using first- through third-passage cultures of BTM cells. For [Ca2+]i and shape change assessment, BTM cells were loaded with fura-2 acetoxymethyl ester, and individual fluorescence images were analyzed after the different experimental manipulations. C studies were performed in vitro using human and bovine anterior segments perfused at constant pressure.


RESULTS
Bradykinin at 10(-6) M elicited a [Ca2+]i increase of 8 to 10 times the basal levels in 90% of the studied cells. From the responder cells, 60% elicited a 15%+/-1% reduction of the initial cell area, and 37% showed a 13%+/-2% reduction of their major axis. Bradykinin failed to induce any effect in the presence of the BK-B2 receptor antagonist HOE-140. Zero [Ca2+]o the depletion of intracellular stores with thapsigargin, or the presence of the calmodulin antagonist W13, decreased the BK response significantly (P < 0.001; P < 0.001; and P < 0.05). A second application of BK elicited a significantly lower (P < 0.001) response than the previous one. Perfusion with 10(-6) M BK decreased CD, calculated as the area under the curve, by 13%+/-4% (P < 0.05) in human anterior segments and 12%+/-4% (P < 0.05) in bovine anterior segments. The presence of 10(-6) M HOE-140, a BK-B2 receptor antagonist, completely blocked the decrease in C after perfusion with BK.


CONCLUSIONS
The C of human and bovine trabecular meshwork (perfused anterior segments) is decreased by BK, acting through BK-B2 receptors. Primary cultured BTM cells respond to BK stimulation by increasing their [Ca2+]i by mobilization of extracellular and intracellular Ca2+. Moreover, these cells are reduced in area and their major axis shortened after the [Ca2+]i peak elicited by BK through BK-B2 receptors. The [Ca2+]i mobilization and shape changes are calmodulin dependent. Taking into account the [Ca2+]i mobilization, the BTM shape changes, the decrease of C, and the temporal sequence of these parameters, a contraction of trabecular meshwork cells related to the functional role of trabecular meshwork is discussed.
Llobet
A
A
Departament de Ciències Fisiològiques, Hospital Clínic, Institut d'Investigacions Biomèdiques August Pi i Sunyer, Universitat de Barcelona, Spain.
Gual
A
A
Palés
J
J
Barraquer
R
R
Tobías
E
E
Nicolás
J M
JM
D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D001082:Aqueous Humor / Q000378:metabolism*; D001920:Bradykinin / Q000031:analogs & derivatives / Q000494:pharmacology*; D065168:Bradykinin Receptor Antagonists; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000037:antagonists & inhibitors; D002417:Cattle; D048429:Cell Size / Q000187:drug effects; D002478:Cells, Cultured; D016257:Fura-2 / Q000378:metabolism; D006801:Humans; D010477:Perfusion; D013449:Sulfonamides / Q000494:pharmacology; D014129:Trabecular Meshwork / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism
D065168:Bradykinin Receptor Antagonists; D002147:Calmodulin; D013449:Sulfonamides; C065679:icatibant; C033772:N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide; D001920:Bradykinin; D002118:Calcium; D016257:Fura-2