Regional differences in functional receptor distribution and calcium mobilization in the intact human lens

PURPOSE. To investigate regional differences in Ca2+ mobilization kinetics in the intact human lens produced by exposure to agonists of tyrosine-kinas e and G-protein-coupled receptors and to characterize the major receptor su btypes involved in Ca2+ signaling in the different regions. METHODS. Whole human lenses were placed anterior side down in a plastic cha mber and perifused with artificial aqueous humor (AAH) at 30 degreesC. Afte r fura-2 incorporation, cytosolic Ca2+ levels were monitored by using epifl uorescence techniques in either the equatorial or central anterior epitheli al cells of the intact lens. Agonists dissolved in AAH were applied to the lens in successive short pulses. RESULTS. Central anterior lens epithelial cells produced a large response t o 10 muM acetylcholine (ACh) and histamine; only a small response to adenos ine triphosphate (ATP); and no response to 10 muM adrenalin, 10 ng/ml epith elial growth factor (EGF) or TGF alpha, or 50 ng/ml platelet-derived growth factor (PDGF)-AB. Conversely, the equatorial cells produced a strong respo nse to 10 muM ATP and histamine, 10 ng/ml EGF (or TGFa), and 50 ng/ml PDGF- AB, but failed to respond to 10 AM ACh or 10 muM adrenalin. The EGF-induced response in the equatorial cells was blocked completely by tyrphostin (AG1 478), a specific inhibitor of the EGF receptor tyrosine kinase. Carbachol, a nonhydrolyzable analogue of ACh, and pilocarpine, the MI muscarinic recep tor-specific agonist, both produced the same trend of response amplitude el icited by ACh in each region of the lens. The potency order of purinergic a gonist-induced Ca2+ mobilization at the equator was consistent with the P2Y (2) receptor subtype. The histamine-induced response was abolished by 10 mu M triprolidine, a specific H-1 receptor antagonist, but remained unaffected by the specific H-2 and H-3 antagonists, ranitidine and thioperamide, resp ectively. CONCLUSIONS. There is a spatial heterogeneity in functional receptor activi ty in different regions of the whole lens. The important growth factor rece ptors for EGF and PDGF are functionally active only in the equatorial cells of the mature human lens. This study further shows that the ACh, histamine , and ATP-induced responses arise from the activation of Ml muscarinic, H-1 histamine, and P2Y(2) purinergic receptors, respectively.