Bleached pigment produces a maintained decrease in outer segment Ca2+ in salamander rods.
A spot confocal microscope based on an argon ion laser was used to make measurements of cytoplasmic calcium concentration (Ca2+i) from the outer segment of an isolated rod loaded with the fluorescent calcium indicator fluo-3 during simultaneous suction pipette recording of the photoresponse. The decline in fluo-3 fluorescence from a rod exposed to saturating illumination was best fitted by two exponentials of approximately equal amplitude with time constants of 260 and 2,200 ms. Calibration of fluo-3 fluorescence in situ yielded Ca2+i estimates of 670 +/- 250 nM in a dark-adapted rod and 30 +/- 10 nM during response saturation after exposure to bright light (mean +/- SD). The resting level of Ca2+i was significantly reduced after bleaching by the laser spot, peak fluo-3 fluorescence falling to 56 +/- 5% (SEM, n = 9) of its value in the dark-adapted rod. Regeneration of the photopigment with exogenous 11-cis-retinal restored peak fluo-3 fluorescence to a value not significantly different from that originally measured in darkness, indicating restoration of the dark-adapted level of Ca2+i. These results are consistent with the notion that sustained activation of the transduction cascade by bleached pigment produces a sustained decrease in rod outer segment Ca2+i, which may be responsible for the bleach-induced adaptation of the kinetics and sensitivity of the photoresponse.
Sampath
A P
AP
Department of Physiological Science, University of California, Los Angeles, Los Angeles, California 90095, USA.
Matthews
H R
HR
Cornwall
M C
MC
Fain
G L
GL
D000221:Adaptation, Ocular / Q000502:physiology; D000557:Ambystoma; D000814:Aniline Compounds; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D003623:Dark Adaptation / Q000502:physiology; D005456:Fluorescent Dyes; D007700:Kinetics; D018613:Microscopy, Confocal; D012168:Retinal Pigments / Q000378:metabolism; D012374:Rod Cell Outer Segment / Q000378:metabolism*; D014785:Vision, Ocular / Q000502:physiology*; D014966:Xanthenes
D000814:Aniline Compounds; D005456:Fluorescent Dyes; D012168:Retinal Pigments; D014966:Xanthenes; C059715:Fluo-3; D002118:Calcium