Dihydropyridine‐sensitive low‐threshold calcium channels in isolated rat hypothalamic neurones.

Abstract

1. Low-voltage-activated Ca2+ channels which produce a transient inward current were studied in neurones freshly isolated from the ventromedial hypothalamic regions of the rat. Membrane currents were recorded using a suction-pipette technique which allows for internal perfusion under a single-electrode voltage clamp. A concentration jump technique was also used for rapid drug application. 2. In most cells superfused to potentials more positive than -65 mV from a holding potential of -100 mV. Such a low-threshold Ca2+ current could easily be separated from a high-threshold, steady type of Ca2+ current by selecting the holding and test potential levels, as well as by resistance to the wash-out during cell dialysis. 3. Activation and inactivation processes of the low-threshold Ca2+ current were highly potential dependent at 20-22.degree.C. For a test potential change from -60 to +20 mV, the time to peak of the current decreased from 45 to 9 ms, and the time constant of the current decay decreased from 90 to 40 ms. The steady-state inactivation occurred at very negative potentials, reaching a 50% level at -93 mV. Recovery from inactivation showed a time constant between 2.63 and 0.94 s for a potential changes from -80 to -120 mV. 4. The amplitude of the low-threshold Ca2+ current depended on the external Ca2+ concentration ([Ca2+]0), approaching saturation at 100 mM [Ca2+]0. Ba2+ substituted for Ca2+ reduced the current amplitude by 30-50% while Sr2+ produced no definite changes in the current amplitude. 5. The low-threshold Ca2+ current was blocked by various di- or trivalent cations in the sequence of La3+ > Zn2+ > Cd2+ > Ni2+ > Co2+. The corresponding apparent dissociation constants (KD) were 7 .times. 10-7, 1 .times. 10-4, 3 .times. 10-4, , 6 .times. 10-4 and 3 .times. 10-3 M. 6. Various organic Ca2+ antagonists were effective in blocking the low-threshold Ca2+ current in the following sequences: flunarizine > nicardipine > nifedipine > nimodipine > D600 (methoxyverapamil) > diltiazem. The corresponding KDs were 7 .times. 10-7, 3.5 .times. 10-6, 5 .times. 10-6, 7 .times. 10-6, 5 .times. 10-5 and 7 .times. 10-5M. These Ca2+ antagonists induced a use-dependent decrease in the current amplitude. Bay K 8644 blocked the current in a manner similar to that seen with other dihydropyridine derivatives. We conclude from the study that pharmacological properties of low-threshold Ca2+ channels in CNS neurones differ from those in peripheral neurones.