The Journal of Neuroscience, April 1, 2003, 23(7):2706
Exocytosis at the Ribbon Synapse of Retinal Bipolar Cells Studied
in Patches of Presynaptic Membrane
Artur
Llobet,
Anne
Cooke, and
Leon
Lagnado
Medical Research Council Laboratory of Molecular Biology, Cambridge
CB2 2QH, United Kingdom
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
Ca2+ influx or loading the terminal with EGTA. The
increase in capacitance declined as the depolarization approached the
reversal potential for Ca2+, indicating that it was
a result of Ca2+-dependent exocytosis. The
conductance increase was caused by KCa channels that were
also activated by Ca2+ influx. Two observations
indicated that sites of exocytosis and endocytosis colocalized with
clusters of Ca2+ channels and KCa
channels; the initial rate of exocytosis was correlated with the
activation of KCa channels, and exocytosis did not occur in
the 41% of patches lacking this conductance. Electron microscopy
demonstrated that there were ~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/µm2. The density of ribbons was 0.10 ± 0.02/µm2, predicting that ~43% of cell-attached
patches would lack an active zone. The density of
Ca2+ channel clusters assayed by capacitance and
conductance responses was therefore similar to the density of ribbons.
These results are consistent with the idea that Ca2+
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
Ca2+ from Ca2+ channel clusters.
Key words:
exocytosis; synaptic terminal; capacitance; calcium; vesicle; endocytosis
Copyright © 2003 Society for Neuroscience 0270-6474/03/2372706-09$05.00/0
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