Ca2+-dependent protein kinase injection in a photoreceptor mimics biophysical effects of associative learning.

Iontophoretic injection of phosphorylase kinase, a Ca2+-calmodulin-dependent protein kinase, increased input resistance, enhanced the long-lasting depolarization component of the light response, and reduced the early transient outward K+ current, IA, and the late K+ currents, IB, in type B photoreceptors of Hermissenda crassicornis in a Ca2+-dependent manner. Since behavioral and biophysical studies have shown that similar membrane changes persist after associative conditioning, these results suggest that Ca2+-dependent protein phosphorylation could mediate the long-term modulation of specific K+ channels as a step in the generation of a coditioned behavioral change.

Protein kinase injection reduces voltage-dependent potassium currents.

Intracellular iontophoretic injection of the catalytic subunit of cyclic adenosine monophosphate-dependent protein kinase increased input resistance and decreased a delayed voltage-dependent K+ current of the type B photoreceptor in the nudibranch Hermissenda crassicornis to a greater extent than an early, rapidly inactivating K+ current (IA). This injection also enhanced the long-lasting depolarization of type B cells after a light step. These findings suggest the involvement of cyclic adenosine monophosphate-dependent phosphorylation in the differential regulation of photoreceptor K+ currents particularly during illumination. On the other hand, conditioning-induced changes in IA may also be regulated by a different type of phosphorylation (for example, Ca2+-dependent).

Double-blind study of pulsing magnetic field effects on multiple sclerosis.

We performed a double-blind study to measure the clinical and subclinical effects of an alternative medicine magnetic device on disease activity in multiple sclerosis (MS). The MS patients were exposed to a magnetic pulsing device (Enermed) where the frequency of the magnetic pulse was in the 4-13 Hz range (50-100 milliGauss). A total of 30 MS patients wore the device on preselected sites between 10 and 24 hours a day for 2 months. Half of the patients (15) randomly received an Enermed device that was magnetically inactive and the other half received an active device. Each MS patient received a set of tests to evaluate MS disease status before and after wearing the Enermed device. The tests included (1) a clinical rating (Kurtzke, EDSS), (2) patient-reported performance scales, and (3) quantitative electroencephalography (QEEG) during a language task. Although there was no significant change between pretreatment and posttreatment in the EDSS scale, there was a significant improvement in the performance scale (PS) combined rating for bladder control, cognitive function, fatigue level, mobility, spasticity, and vision (active group -3.83 +/- 1.08, p < 0.005; placebo group -0.17 +/- 1.07, change in PS scale). There was also a significant change between pretreatment and posttreatment in alpha EEG magnitude during the language task recorded at various electrode sites on the left side. In this double-blind, placebo-controlled study, we have demonstrated a statistically significant effect of the Enermed magnetic pulsing device on patient performance scales and on alpha EEG magnitude during a language task.

Modulation of calcium current and diverse K+ currents in identified Hermissenda neurons by small cardioactive peptide B.

The molluscan neuropeptides, small cardioactive peptides A and B (SCPA,B), are known to modulate the responses of many molluscan central and peripheral target cells (see review by Lloyd, 1986), but their full range of physiological actions remains unknown. External application of SCPB (1-10 microM) modified diverse ionic conductances in a set of giant identifiable neurons in the brain of the marine mollusk Hermissenda crassicornis. SCPB caused a transient depolarization and increased input resistance that enhanced or promoted cell firing. Under voltage-clamp, SCPB reduced a "background" residual current (IR), reduced early transient K+ current (IA), reduced a delayed K+ current (IK(V], and enhanced ICa, IBa, and a Ca2+-activated K+ current, IK(Ca). In tetraethylammonium chloride (TEA) saline, SCPB enhanced the amplitude and duration and reduced the threshold of evoked Ca and Ba spikes. Immunocytochemical staining techniques have localized an endogenous SCPB-like peptide in numerous somata and their neurites in the nervous system of Hermissenda (Longley and Longley, 1985; Watson and Willows, 1986). These data are consistent with a role for SCPB as a neurotransmitter/neurohormone modulator of neuronal excitability in Hermissenda. A neurotransmitter role for endogenous SCPs has been proposed for a synaptic pair of cultured neurons in the Aplysia buccal ganglion (Lloyd et al., 1986). SCPB has been implicated in the control of feeding motor output in Aplysia (Sossin et al., 1986) and Tritonia (Willows and Watson, 1986), and in the presynaptic facilitation of sensory neurons mediating the gill and siphon defensive withdrawal reflex in Aplysia (Abrams et al., 1984).

Characterization of voltage-activated currents in Hermissenda type B-photoreceptors.

Two distinct voltage-dependent K+ currents have been previously identified in the type B-photoreceptors of Hermissenda: an early, rapidly inactivating K+ current (IA), and a late, 4-AP- and TEA-resistant voltage and Ca(2+)-dependent current (IK(Ca)). Using conventional two-electrode voltage-clamp techniques, we have characterized two additional currents, a late voltage-dependent outward K+ current (IK(v)) and a voltage-dependent inward current identified as an inward rectifier (Iir). In addition, we have further studied the activation-inactivation kinetics of IA. In 0-Na+ ASW, Iir activates at a potential of > -50 mV, is steeply voltage-dependent and noninactivating, and reaches steady-state within 800 msec to 3 sec at -100 mV. In addition to the variability in activation kinetics, there was also considerable variability in Iir magnitude (-5 to -80 nA, at -100 mV). Iir was blocked by external 4-AP (5 mM), external and internal TEA, internal Cs+, but not external Ba2+. The major component of outward K+ current in type B-photoreceptors is IK(v), the delayed rectifier. IK(v) was isolated after removal of IA and IK(Ca). IK(v) activates at around -25 mV or more positive membrane potentials and its activation and inactivation are strongly voltage dependent. IK(v) inactivation to steady state is reached within 1.5-2.5 sec. The wide range of activation-inactivation rates suggests that there may be kinetic subtypes of IK(v). The proposed "slow" IK(v) peaked in 50-90 msec at +30 mV, and decayed with a single exponential component with an average tau off of 279 msec. Proposed "intermediate" and "fast" IK(v) subtypes peaked within 12-50 msec at +30 mV, and had a decay fitted by two exponentials, with an average tau 1 of 147 msec and tau 2 of 275 msec, respectively. IK(v) exhibited marked twin-pulse inactivation with a recovery time of 30-40 sec, and also exhibited time- and voltage-dependent cumulative inactivation to repeated depolarizing pulses. Both types of inactivation were quickly removed by a prepulse hyperpolarization. 4-AP (5 mM) produced partial to complete block of the inactivating component of IK(v), leaving only a residual sustained component. Complete block of the transient and sustained components of IK(v) was obtained by 100 mM TEA. Reliable voltage separation of IA from IK(v) was achieved by activating IA in the range of -50 to -20 mV, from a Vh of -80 mV. Voltage-dependent steady-state inactivation curves for IA were determined, yielding an average h0.5 value of -56 mV.(ABSTRACT TRUNCATED AT 250 WORDS)

Soma spike of neuroendocrine bag cells of Aplysia californica.

Soma action potentials of the neuroendocrine bag cells of Aplysia californica were studied with intracellular recording and current injection. Spikes in artificial sea water (ASW) were either graded with increasing depolarizing current pulses, or had a well-defined threshold. The latter spikes typically had faster rise times with larger overshoots and hyperpolarizing afterpotentials. Repetitive stimulation led to spike potentiation (SP), manifested as an increase in overshoot amplitude and duration of successive spikes in a train. SP was usually detectable at 0.5 Hz, and maximal between 0.8 and 4 Hz. Concomitant accommodation occurred rapidly at greater than or equal to 5 Hz. The increase in spike duration during SP resulted from a progressive enhancement of an inflection on the repolarizing phase. The inflection was dependent on membrane potential; small depolarizations (5-10 mV) enhanced it; hyperpolarization (less than 35 mV) reduced it. Solutions with O--Na+ (Tris-substituted) or O--Ca2+ (1 mM EGTA) revealed mixed Na+/Ca2+ spikes with variable degrees of Na+ versus Ca2+ dominance. Cd2+, Co2+, and Mn2+ reversibly abolished the inflection on the repolarizing phase, indicating that it is Ca2+ mediated; the spike was reduced irreversibly at higher concentrations. SP was generally reduced only if the spike was severely attenuated. It is proposed that SP results primarily from a voltage- and time-dependent potassium inactivation which then unmasks a calcium current. SP may play a role in augmenting the release of egg-laying hormone.

Serotonin decreases a background current and increases calcium and calcium-activated current in pedal neurons of Hermissenda.

The effects of serotonin (5-HT) on membrane potential, membrane resistance, and select ionic currents were examined in large pedal neurons (LP1, LP3) of the mollusk Hermissenda. Calcium (Ca) action potentials were evoked in sodium-free artificial seawater containing tetramethylammonium, tetraethylammonium, and 4-aminopyridine (0-Na, 4-AP, TEA ASW). They failed at stimulation rates greater than 0.5/sec and were blocked by cadmium (Cd). Under voltage clamp the calcium current (ICa) responsible for them also failed with repeated stimulation. Thus, ICa inactivation accounts for refractoriness of the Ca action potential. The addition of 10 microM 5-HT to 0-Na, 4-AP, TEA ASW produced a slight depolarization and increased excitability and input resistance. Under voltage clamp the background current decreased. The voltage-dependent inward, late outward, and outward tail currents, sensitive to Cd, increased. ICa inactivation persisted. Under voltage clamp with Ca influx blocked by Cd, the addition of 10 microM 5-HT decreased the remaining current uniformly over membrane potentials of -10 to -100 mV. Thus, 5-HT reduces a background current that is active within the physiological range of the membrane potential, voltage insensitive, independent of Ca influx, noninactivating, and not blocked by 4-AP or TEA.

Differential modulation of voltage-dependent currents in Hermissenda type B photoreceptors by serotonin.

1. In identified photoreceptors in the eyes of Hermissenda, serotonin (5-HT) enhances the peak and plateau phases of light-evoked generator potentials and modulates light-dependent and voltage-dependent currents. In addition, electrophysiological and morphological studies indicate that 5-HT may contribute to cellular plasticity detected in the visual system of Hermissenda produced by classical conditioning procedures. With the use of conventional two-electrode voltage-clamp techniques, we examined the effects of 5-HT on three distinct currents recorded across the soma membranes of identified lateral and medial type B photoreceptors in the isolated circumesophageal nervous system. 2. The inward rectifier (Iir), a putative K(+)-dependent conductance, activates in 0-Na artificial seawater at membrane potentials greater than -60 mV, is voltage dependent, noninactivating, and reaches steady-state within 800 ms to 3 s at -100 mV. Bath application of 10(-4) M 5-HT consistently enhanced the magnitude of Iir at all potentials tested (-60 to -100 mV) and, in some cases, allowed expression of Iir, which was not initially detectable before the application of 5-HT. 3. The major component of outward K+ current in type B photoreceptors with IA and IK(Ca) blocked is the delayed rectifier [IK(v)]. 5-HT (10(-4) M) produced both an increase as well as a decrease in the peak amplitude of IK(v) and consistently slowed its inactivation rate and reduced twin-pulse inactivation. 4. A previously identified outward K+ current in type B photoreceptors has properties that are similar to the A current (IA).(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of delta9-tetrahydrocannabinol on action potentials in the mollusc Aplysia.

The effects of delta9-tetrahydrocannabinol (delta9-THC) on action potentials were examined during intrasomatic recordings from the isolated buccal and parieto-visceral ganglia of Aplysia californica. When added to the saline solution bathing the preparation, the compound (in concentrations 10(-4) - 10(-5) M) caused a reduction in spike overshoot (15-20% of total amplitude) and increased the lability of responses to electrical stimulation. The somatic membrane appeared to be more affected than the axonal membrane. Diffusion barriers in the ganglion probably account for the high degree of variability in drug response, such that both of the characteristic changes were observed in only about 30% of the tests. This is the first report to describe effects of delta9-THC on invertebrate neurones. The results indicate that delta9-THC causes a depression in nerve cell excitability, and these data are consistent with reported effects of THC compounds in mammals.

Serotonin differentially modulates two K+ currents in the Retzius cell of the leech.

The effects of 100 mumol l-1 serotonin (5-HT) were investigated on the Na+- and Ca2+-dependent action potential and distinct K+ currents in the Retzius (R) cells of the hirudinid leeches Macrobdella decora and Hirudo medicinalis by conventional current-clamp and two-microelectrode voltage-clamp techniques. 1. In normal Na+-containing Ringer, 5-HT decreased the duration of the action potential prolonged by 5 mmol l-1 tetraethylammonium (TEA+) chloride. 2. In Na+-free saline containing 25 mumol l-1 TEA+ to block IK, 5-HT reduced the amplitude and duration of Ca2+ spikes evoked by intracellular current injection. 3. Under voltage-clamp, 5-HT enhanced the peak amplitude of an early transient 4-aminopyridine (4-AP)-sensitive, voltage-dependent outward current, termed IA. A small but significant increase in the time constant of inactivation (tau off) of IA was also measured after exposure to 5-HT. 4. 5-HT suppressed the peak and steady-state amplitudes of a delayed TEA+-sensitive, voltage-dependent outward current, termed IK. These results demonstrate differential simultaneous modulation of distinct K+ currents in the Retzius cell of the leech by the endogenous transmitter serotonin. These cells contain and release 5-HT, and are believed to be multifunction neurons implicated in feeding and swimming. This modulation may change the excitable properties of the cell, leading to a negative feedback autoregulation of its transmitter output.

Presynaptic spike broadening reduces junctional potential amplitude.

Presynaptic modulation of action potential duration may regulate synaptic transmission in both vertebrates and invertebrates. Such synaptic plasticity is brought about by modifications to membrane currents at presynaptic release sites, which, in turn, lead to changes in the concentration of cytosolic calcium available for mediating transmitter release. The 'primitive' neuromuscular junction of the jellyfish Polyorchis penicillatus is a useful model of presynaptic modulation. In this study, we show that the durations of action potentials in the motor neurons of this jellyfish are negatively correlated with the amplitude of excitatory junctional potentials. We present data from in vitro voltage-clamp experiments showing that short duration voltage spikes, which elicit large excitatory junctional potentials in vivo, produce larger and briefer calcium currents than do long duration action potentials, which elicit small excitatory junctional potentials.