Exocytosis at the ribbon synapse of retinal bipolar cells studied in patches of presynaptic membrane.
The distribution of exocytic sites and ion channels in the synaptic terminal of retinal bipolar cells was investigated by measuring capacitance and conductance changes in cell-attached patches of presynaptic membrane. Patch depolarization evoked capacitance and conductance increases that were inhibited by blocking Ca(2+) influx or loading the terminal with EGTA. The increase in capacitance declined as the depolarization approached the reversal potential for Ca(2+), indicating that it was a result of Ca(2+)-dependent exocytosis. The conductance increase was caused by K(Ca) channels that were also activated by Ca(2+) influx. Two observations indicated that sites of exocytosis and endocytosis colocalized with clusters of Ca(2+) channels and K(Ca) channels; the initial rate of exocytosis was correlated with the activation of K(Ca) channels, and exocytosis did not occur in the 41% of patches lacking this conductance. Electron microscopy demonstrated that there were approximately 16 vesicles docked to the plasma membrane at each active zone marked by a ribbon, but vesicles were also attached to the rest of the membrane at a density of 1.5/microm(2). The density of ribbons was 0.10 +/- 0.02/microm(2), predicting that approximately 43% of cell-attached patches would lack an active zone. The density of Ca(2+) channel clusters assayed by capacitance and conductance responses was therefore similar to the density of ribbons. These results are consistent with the idea that Ca(2+) channel clusters were colocalized with ribbons but do not exclude the possibility that calcium channels also occurred at other sites. The wide distribution of vesicles docked to the plasma membrane suggests that exocytosis might also be triggered by the spread of Ca(2+) from Ca(2+) channel clusters.
Llobet
Artur
A
Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 2QH, United Kingdom.
Cooke
Anne
A
Lagnado
Leon
L
D000818:Animals; D015220:Calcium Channels / Q000032:analysis; D002478:Cells, Cultured; D004553:Electric Conductivity; D004705:Endocytosis; D005089:Exocytosis; D006054:Goldfish; D007700:Kinetics; D009474:Neurons / Q000502:physiology*; D018408:Patch-Clamp Techniques; D024681:Potassium Channels, Calcium-Activated / Q000032:analysis; D017729:Presynaptic Terminals / Q000737:chemistry / Q000502:physiology / Q000648:ultrastructure*; D012160:Retina / Q000166:cytology*; D013570:Synaptic Membranes / Q000737:chemistry / Q000502:physiology / Q000648:ultrastructure*; D013572:Synaptic Vesicles / Q000648:ultrastructure
D015220:Calcium Channels; D024681:Potassium Channels, Calcium-Activated