Norio Akaike

Actions of verapamil, diltiazem and other divalent cations on the calcium-current of Helix neurones

1 Effects of organic Ca2+‐antagonists, verapamil and diltiazem, and cations, Ni2+, Mn2+, Co2+ and La3+ on Ca2+ current (ICa) separated from other ionic currents in a Helix neurone were studied. A suction pipette technique which allows internal perfusion of the cell body and voltage clamp was used 2 Verapamil and diltiazem (10−6‐10−4m) increased the threshold, and decreased both the amplitude and rate of rise of the soma Ca2+‐spike. Both agents inhibited ICa over the entire range of the current‐voltage (I‐V) relationship dose‐dependently, without shifting the threshold of the I‐V relationship. Increases in external Ca2+ overcame the inhibitory action of the agents 3 Divalent cations, Ni2+, Mn2+, Co2+ and the trivalent cation, La3+ inhibited ICa dose‐dependently, but induced shifts of the I‐V relationship to more positive voltages. The order of potency of inhibition of ICa among these cations was as follows; Ni2+ > La3+ > Mn2+ > Co2+ 4 Double reciprocal plots for peak ICa versus external Ca2+ concentrations in the presence or absence of both organic and inorganic Ca2+‐antagonists intersect at the ordinate. Results indicate that both organic and inorganic Ca2+‐antagonists compete for Ca2+ at the common binding site for Ca2+ 5 Internal application of the organic Ca2+‐antagonists (10−4m) inhibited ICa in a time‐dependent manner to about 40–60% of the control. Ni2+, when applied internally, also depressed ICa 6 The results provide evidence that organic Ca2+‐antagonists occupy the binding site for Ca2+ in a competitive manner at the surface of the soma membrane of the Helix neurone, while divalent and trivalent cations, in addition to inhibiting ICa in a similar manner to the organic Ca2+‐antagonists, change the surface charge of the soma membrane.

FURTHER ANALYSIS OF INHIBITORY EFFECTS OF PROPRANOLOL AND LOCAL ANAESTHETICS ON THE CALCIUM CURRENT IN Helix NEURONES

1 The effects of propranolol and local anaesthetics on Ca2+ current (ICa), individually separated from other ionic currents, in Helix neurones were studied under voltage clamp, using a suction pipette technique. 2 Increases in external Ca2+ concentrations overcame the inhibitory action of propranolol on lea‐Double reciprocal plots for peak ICa versus external Ca2+ concentrations in the presence or absence of propranolol did not intersect at the ordinate. 3 Internal application of propranolol (10−4m) inhibited ICa to about 40–60% of the control in a time‐dependent manner. 4 Lignocaine and procaine at concentrations of 10−3–10−2m inhibited ICa without shifting the threshold in the I‐V relationships. Internal application of lignocaine (10−3–10−2 m) also inhibited ICa: the ratio of depression of the ICa was almost equivalent to that of the agent applied externally. 5 The results provide evidence that propranolol inhibits ICa in a noncompetitive manner with Ca2+ at the cell membrane, and suggest that the agents may occupy the receptor site in the Ca2+ ‐channel somewhere between the outer surface and inner phase of the membrane.

INHIBITORY EFFECTS OF PROPRANOLOL ON THE CALCIUM CURRENT OF Helix NEURONES

1 Calcium current (ICa) and potassium current (IK) in isolated nerve cell bodies of Helix aspersa were independently separated after suppression of Na+ and K+ currents, and Na+ and Ca2+ currents, respectively. The suction pipette method was used. Under voltage clamp conditions, the effects of propranolol on both the ICa and IK were examined. 2 Propranolol inhibits the ICa at relatively low concentrations (10−6‐10−5m). The inhibitory action was dose‐dependent. 3 The IK was moderately suppressed by the drug at high concentrations (10−5‐5 × 10−4m). 4 Results provide evidence for a new aspect of the pharmacological action of propranolol on the excitable cell membrane.

Role of anions and cations in frog taste cell stimulation.

Abstract 1. 1. Role of cations and anions in stimulating frog taste receptors was investigated by recording changes in the membrane potential and the effective resistance in taste cells with a glass capillary microelectrode. 2. 2. The results indicate that the membrane potential in taste cells originates from the diffusion potential due to receptor membrane permeability to monovalent cations and anions, and that anions, in the order of the lyotropic series, modify the membrane permeability to both ion species. 3. 3. Depolarizations by Ca and other divalent cations in taste cells and their inhibition by anions were also examined, and their mechanisms were discussed.

Effects of n-alkanols on the calcium current of intracellularly perfused neurons of Helix aspersa

The effects of n-alkanols on the calcium current (ICa) were studied in molluscan neurons perfused intracellularly and voltage clamped using a suction pipette technique. All n-alkanols employed in this experiment (methanol, ethanol and butanol) decreased the peak amplitude of ICa and caused acceleration of the decay of ICa in a dose-dependent manner at all membrane potentials. The concentrations of n-alkanols required for these actions decreased as the hydrocarbon chain increased in length. The results suggest that these effects on the ICa of molluscan neurons may be related to the lipophilic properties of n-alkanols.

Water response in frog taste cells

Abstract o 1. Glossopharyngeal nerve responses to simulations of three different portions of the tongue by water and various chemicals were recorded. Water elicited a slowly rising and persistent response, but no difference in sensitivity to water was found among the three portions. 2. Water produced a hyperpolarization in taste cells, but the hyperpolarization decreased by about 10–40 mV during water stimulation. The decrease of the membrane potential from the hyperpolarized level has been considered to be a factor that initiates impulses at the sensory nerve terminal.

CNS effects on muscle Na/K levels in hypokalemia

Diets deficient in K + are known to reduce the K + concentration of the blood plasma and skeletal muscles while having little or no effect on K + levels in the cerebrospinal fluid or brain tissue1, 2. I t has been proposed that the choroid plexus contributes to brain homeostasis by maintaining a constant level of K + in the cerebrospinal fluid 1. The reduced K + content of muscle during hypokalemia might be attributed to a partial inhibition of the N a K pump due to the reduced plasma K + (see refs. 1 and 2). However, in rats fed potassium-deficient diets for 3-4 weeks plasma K + levels do not fall much below 2.5 mM (see ref. 3). The muscles from these rats show significant increases in intraceUular Na + ([Na]i) levels and decreases in intracellular K + ([K]i). When these muscles are excised and placed in a Krebs solution containing 2.5 mM K + at 37 ~C, the [Na]i level is promptly reduced while the [K]i level is increased 3,4. Therefore, the high [Na]i and low [K]i levels obtained from in situ muscles of potassium-deficient rats are not the direct result of pump inhibition by low plasma K + levels. Soleus muscles (SOLs) from male, Wistar rats were used. The hypokalemic group was fed a potassium-deficient diet for 3-4 weeks. All rats were in good health at the time of the experiments. In some animals, nerves were sectioned as described below under ether anesthesia. The Na + and K + contents of muscles and serum samples were measured with a flame spectrophotometer. The method used for the determination of [Na]i and [K]i is identical to that described elsewhere 4. In the rats fed the potassium-deficient diet for 3-4 weeks, there was a significant decrease in both body weight (P < 0.01) and plasma K + levels (P < 0.001), but no significant changes in plasma N a ÷ levels. During the potassium-deficient period, the decrease in muscle K + content is accompanied by an almost equivalent increase in muscle Na + content. Unilateral denervation of SOLs was made by section of the tibial nerve branch in potassium-deprived rats under ether anesthesia. The contralateral nerve was shamoperated. Both muscles of each pair were dissected out at various intervals following the surgical procedure and prepared for Na + and K + analysis. A dramatic effect of

Intracellular ion concentration and electrical activity in potassium-depleted mammalian soleus muscle fibers.

SummaryThe relationship between the intracellular cation concentration and the membrane potential has been studied in ‘Na-rich’ soleus muscle fibers of rats which had been fed a K-free diet for 10–50 days. The resting potentials of ‘Na-rich’ muscle fibers closely agreed with the theoretical potential expected from ionic theory when a quantitative dissociation of active cation transport with Na ions extrusion exceeding K ions uptake was eliminated due to the recovery of ‘Na-rich’ fibers in Krebs solution with 10 mM K for 2 h at 37°C. The hyperpolarized membrane potentials during cellular Na ions extrusion were accounted for by the sum of the potentials produced by the electrogenic Na-pump and by the ionic diffusion potential. On the other hand, the amplitude of overshoot of action potentials decreased linearly with the logarithmic increase of the intracellular Na concentration ([Na]i). The maximum rate of rise of action potentials also changed as a function of [Na]i, though, at the early period of K-deficiency the inhibitory effect of the increased [Na]i on the maximum rate of rise was transiently masked by the hyperpolarization produced by the electrogenic Na-pump which secondarily led to a progressive reduction of Na inactivation, while the maximum rate of fall was a linear function of [K]i.

Activation of electrogenic sodium pump in mammalian skeletal muscle by external cations

SummaryThe effect of change of the external ionic composition on “Na-loaded” and “K-depleted” soleus muscle fibres of K-deficient rats was investigated by recording resting membrane potentials. The addition of K, Rb, Cs and NH4 ions to K-free Krebs solution bathing “Na-rich” muscles resulted in a rapid hyperpolarization. The hyperpolarization was abolished by removing the above cations, cooling to ca. 4° C, and adding 0.1 mM ouabain. The effectiveness of cations for activating the electrogenic Na pump was Rb≧K>NH4>Cs, and NH4 ions seemed to be unique in their stimulating action. The resting cell membrane of “Na-rich” muscles is permeable to cations in the order of Rb=K>Cs>NH4. Reducing Na ions in Krebs solution had no effect on the rate of Na-pumping in “Na-rich” muscle fibres at a given K concentration. It is concluded that the external K ions could be replaced by Rb, Cs and NH4 ions in activating the electrogenic Na pump in “Na-rich” soleus muscle fibres, but that the electrogenic Na pump in this tissue does not require the external Na ions.

Operation of an electrogenic Na-pump in mammalian red muscle fibre

Abstract The resting membrane potential in ‘Na-rich’ soleus muscle fibres, obtained from the K-depleted rats fed a K-free diet, was rapidly hyperpolarized beyond the theoretical value derived from the ionic theory or even beyond the potential measured in ‘fresh’ muscle fibres, when 2.5 to 15 mM-K was added to the bathing solution at 37°C. The K-sensitive hyperpolarization was abolished after cooling to ca. 4°C or adding ouabain. Therefore, the observed membrane potential exceeding the calculated potential during hyperpolarization was attributed to an electrogenic Na-pump.