Characterization of the Ca 2+ Current in Freshly Dissociated Crustacean
Peptidergic Neuronal Somata.
Richmond, Janet E., Emanuele Sher, Ian M. Cooke.
B[acute]ek[acute]esy Laboratory of Neurobiology, University of Hawaii,
Honolulu, Hawaii
96822, Permanent address: CNR Center of Cytopharmacology, Milan, Italy.
APStracts 2:0030N, 1995.
SUMMARY AND CONCLUSIONS
1. Freshly dissociated neuronal somata of the crab ( Cardisoma carnifex ) X-
organ were studied in the whole-cell patch clamp configuration. In order to
characterize the Ca 2+ currents in these somata, recordings were made under
conditions designed to suppress K + and Na + currents. 2. In 52 mM
external Ca 2+ the threshold for activation of Ca 2+ currents was above -40
mV, with peak amplitudes occurring around +10 to +20 mV. The full component of
the current was available for activation at -50 mV since no current increase
was observed when the holding potential was increased to -90 mV. These
characteristics of the current characterize it as a high-voltage activated
(HVA) current. 3. The Ca 2+ current was almost completely (60-90%)
inactivated within 200 ms at maximal current potentials (+10 to +20 mV). The
decay was best described by a double-exponential function with a fast and slow
component of inactivation ( f =12 ms and s =64 ms). Both Sr 2+ and Ba 2+
substitutions reduced the rates of inactivation. 4 . In double-pulse
experiments, plots of variable prepulse potential vs. test pulse current
produced a U-shaped curve with test pulse currents showing maximal
inactivation at potentials which produced maximal Ca 2+ influx during the
prepulse. Tail currents also displayed a U-shaped inactivation curve. The
extent of current-dependent inactivation was sequentially reduced by Sr 2+
and Ba 2+ substitutions. These data suggest that inactivation in crab somata
is predominantly Ca 2+ -dependent. The remaining inactivation of Ba 2+
currents suggests that there is also a component of voltage-dependent
inactivation in the somata. 5. Part of the inactivated Ca 2+ current could
be recovered during short (4-10 ms) hyperpolarizing pulses to -130 mV. The
absolute extent of recovery from inactivation was greatest for currents
carried by Ca 2+ rather than Sr 2+ or Ba 2+ . When voltage-dependent
inactivation was dominant (Ba 2+ currents), the relative amount of current
recovered was greater. The data suggest that hyperpolarizing pulses are more
effective in removing voltage-dependent inactivation, but also allow some
recovery from Ca 2+ -dependent inactivation. 6. In the crab saline, which
contained 24 mM Mg 2+ , the amplitudes of currents carried by 52 mM Ca 2+ , Sr
2+ and Ba 2+ were similar. Removing the Mg 2+ from the saline augmented
both the Ba 2+ and Sr 2+ currents relative to the Ca 2+ current. The dose-
response relationship between Mg 2+ concentration and current amplitude was
compared for 52 mM Ca 2+ , Sr 2+ and Ba 2+ . Mg 2+ blocked Ba 2+ >Sr 2+
>Ca 2+ . The ability of Mg 2+ to suppress HVA currents was also dependent
on the concentration of permeant divalent ions used. 7. The ability of
several known inorganic Ca 2+ channel blockers to effect Ca 2+ current
amplitude was determined. The order of blocking potency was La 3+ =Cd 2+ >Ni
2+ =Co 2+ >Mg 2+ . 8. No effects on Ca 2+ current amplitude were found
with nifedepine (10 [mu]M), Bay K 8644 (1 [mu]M), _-conotoxin GVIA, _-Agatoxin
IVA or _-conotoxin MVIIC, indicating that the HVA Ca 2+ current in X-organ
somata is pharmacologically distinct from other characterized channels of the
L-, N-, P-, and Q-types.
Received 12 September 1994; accepted in final form 2 February 1995.
APS Manuscript Number J575-4.
Article publication pending J. Neurophysiol.
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 3 April 1995.