Calcium-activated inward spike after-currents in bursting neurone R15 of Aplysia

Abstract

1. Slow inward and outward after-currents follow action potentials in the bursting pacemaker neurone, R 15, of Aplysia californica. These experiments were performed to examine the role of axo-dendritic calcium influx in activating these after currents. 2. Depolarizing voltage-clamp commands issued at the soma were used to elicit the after-currents. The earlier inward depolarizing after-current of DAC was followed by the hyperpolarizing after-current or HAC. The DAC and HAC appeared at a threshold following depolarizing commands in normal sea water, presumably due to triggering of action potentials in inadequately space-clamped axon. In 100 microM-tetrodotoxin (TTX), the after-currents were graded, increasing gradually in amplitude with increasing voltage or duration of the command. 3. After-current amplitudes varied with the holding potential through the range tested, -40 to -80 mV. DACs were maximum at -40 to -50 mV and decreased in amplitude with hyperpolarization. HACs were maximum at -40 mV and decreased with hyperpolarization to disappear between -70 and -80 mV. 4. The dependence of after-currents upon intracellular calcium accumulation during the depolarizing command was tested in several ways. Bathing R15 in 0 Ca2+-2 mM-EGTA (ethyleneglycol-bis-(beta-aminoethylether)-N,N'-tetraacetic acid) sea water eliminated the after-currents. Bathing in 1 mM-Ca2+ sea water reduced the DAC by 76% and the HAC by 87% following 10 ms long depolarizations to +40 mV. Application of Mn2+ (25 mM) or La3+ (5 mM) blocked the after-currents. Injection of EGTA intracellularly practically eliminated after-currents. Greatly prolonged depolarizations were required to elicit them after EGTA injection. Substitution of Ba2+ for Ca2+ also eliminated after-currents. 5. Sodium-free sea water eliminated the DAC. The HAC following brief (less than 30 ms) depolarizing commands was also eliminated in zero sodium, although longer commands were followed by an outward tail current. 6. Although the after-currents seemed dependent upon calcium influx, they were not suppressed by depolarizing commands whose voltage exceeded the calcium equilibrium potential at the soma as indicated by suppression of the calcium-activated potassium current, or IK(Ca), observed during the depolarization. However, if the extracellular calcium was lowered to 1 mM, large depolarizations did suppress the DAC. 7. Dopamine blocked the after-currents when applied to the axo-dendritic area but not when applied to the soma. Similarly, synaptic inhibition of long duration blocked the after-currents.(ABSTRACT TRUNCATED AT 400 WORDS)