Photoresponses of an extraocular photoreceptor associated with a decrease in membrane conductance in an opisthobranch mollusc

The photoresponse of an extraocular photoreceptor, the photoresponsive neuron (A-P-1) in the abdominal ganglion of Onchidium verruculatum, was studied by using a voltage-clamp with two micropipettes and a monochromatic light. When the A-P-1 was voltage-clamped at resting membrane potential levels, light induced a slowly developing inward current which peaked at about 20 s. A decrease in membrane conductance accompanied this light-induced current which corresponded to the depolarizing photoreceptor potential in the unclamped A-P-1. The relationship between the peak of the current response and light intensity could be predicted by using the modified Michaelis-Menten equation. The spectral sensitivity for the photoresponse had a peak at 490 nm. The steady-state light-induced current was a non-linear function of the membrane potential. The current-voltage relationship for the instantaneous light-induced current was almost linear. In normal (10 mM K+) saline, the polarity of the instantaneous light current reversed from inward to outward at about -67 mV, and doubling the external K+ from 10 to 20 mM shifted the reversal potential to about-50 mV, similar to that predicted by a K+-electrode. These results suggest that the light-induced current or the depolarizing receptor potential of A-P-1 is due to the light suppression of a voltage- and time-dependent K+ current.     

Serotonin involvement during in vitro conditioning of Hermissenda

To determine if serotonin may be involved in associative conditioning-produced changes in the excitability and photoresponses of Type B photoreceptors, isolated nervous systems were exposed to an in vitro conditioning procedure in the presence or absence of drugs that alter normal serotonergic neurotransmission. Pairings of light and intracellular depolarization of a caudal hair cell (in vitro conditioning) produced a pairing-specific depolarization of Type B photoreceptors that was accompanied by an increase in resting input resistance. Treatment of nervous systems with pharmacological agents which disrupt 5-HT neurotransmission attenuated membrane potential and input resistance changes of Type B photoreceptors. These drugs included serotonin uptake inhibitors (imipramine, fluoxetine), a receptor antagonist (bufotenine), and a neurotoxin (5,7-dihydroxytryptamine; 5, 7-DHT). Yohimbine, an alpha 2-receptor antagonist, was without effect. These results, and those in the accompanying papers, suggest that serotonin modulates Type B photoreceptor excitability during associative conditioning.     

Egg-laying hormone, serotonin, and cyclic nucleotide modulation of ionic currents in the identified motoneuron B16 of Aplysia

We have used the 2-electrode voltage-clamp technique to analyze the effects of the neuropeptide, egg-laying hormone (ELH), and the biogenic amine 5-HT on ionic currents in the buccal motoneuron B16 of Aplysia. When B16 is voltage-clamped near resting membrane potential, bath-applied ELH induces a prolonged inward shift in holding current. The ELH-induced inward current is not due to a decrease in the transient, the delayed, or the calcium-activated potassium currents. Current-voltage measurements, along with ion substitution and channel-blocking experiments, indicate that ELH primarily induces or increases a voltage-dependent slow inward current carried by sodium. Serotonin also causes a prolonged inward shift in holding current in B16. Like ELH, 5-HT induces or enhances the voltage-dependent inward current carried by sodium. In addition, 5-HT increases an inwardly rectifying potassium current, and, in some preparations, decreases an outward current that is activated when the cell is depolarized to -40 mV and above. None of the currents modulated by ELH or by 5-HT are affected by 200 microM ouabain or by reducing extracellular chloride concentration. Extracellular application of isobutylmethylxanthine (IBMX), forskolin, 8-bromo-cAMP and 8-bromo-cGMP, and intracellular injection of cAMP elicits the slow inward current carried by sodium. The inward current response to ELH is blocked by prior application of 8-bromo-cAMP, while, under these conditions, 5-HT continues to elicit the increase in inward-rectifier potassium current, but decreases the slow inward sodium current. Serotonin also reduces the slow inward sodium current when applied after ELH. These results suggest that the modulation of B16 by ELH may be mediated entirely by either cAMP or cGMP, while at least a portion of the response to 5-HT may involve an unidentified second messenger in addition to cyclic nucleotides.     

Differential effects of pentylenetetrazole on ion currents of Aplysia neurones

The effect of the convulsant drug pentylenetetrazole (PTZ) on separated membrane current components has been studied in identified voltage-clamped Aplysia neurones. External PTZ blocks the voltage-dependent Na+, Ca2+ currents and the delayed rectifier current (INa, ICa and IK,V, respectively). The amplitude of the Ca2+-activated K+ current (IK,Ca) is increased. The amplitude of the fast inactivating K+ current (IA) is transiently increased at low concentrations of PTZ but is depressed at higher concentrations or after long-lasting application of the drug. The effect of PTZ on leakage current (IL) seems to depend on the cell type. In some cells (R-15, L-7, LP-1) IL is decreased while it is increased in other cells (L-11, BL-1, BR-1). PTZ accelerates the inactivation of IK,V and IA and shifts the current-voltage relation of ICa to negative voltages by 5-8 mV. Pressure injection of PTZ into the neurone did not affect IK,V or IK,Ca. Thus PTZ seems to act on the outside of the plasma membrane. The effect of external PTZ on INa, ICa, IK,V and IL is also observed if the internal Ca2+ activity is buffered with EGTA suggesting that an increase in the internal Ca2+ activity is not involved. At -40 mV PTZ induces a tetrodotoxin-insensitive inward current carried by Na+ ions. PTZ transforms the beating pacemaker cell L-11 into a bursting pacemaker and the bursting pacemaker cell R-15 exhibits 'square-wave'-like oscillations of the membrane potential.     

Different actions of intracellular free calcium on resting and GABA-gated chloride conductances

The effects of voltage-dependent Ca2+ current (ICa) on resting and gamma-aminobutyric acid (GABA)-gated Cl- conductances were studied in isolated frog sensory neurons. The amplitude of GABA-gated Cl- current (ICl) was greatly suppressed by a preceding ICa. The GABA dose-response curve was shifted to the right without changing the maximum response by increasing [Ca2+]i. An ICa-activated ICl was observed as an inward tail current on the 'off' phase of ICa. This tail current was easily saturated by increasing the amount of Ca2+ influx.     

GABA-induced potentiation of neuronal excitability occurs during contiguous pairings with intracellular calcium elevation.

The temporal convergence of neuronal signals is commonly considered as a likely prerequisite for enhanced neuronal excitability underlying the induction of associative memories. Here we report that transmitter application on presynaptic terminals of the Hermissenda Type B photoreceptors, when paired with depolarization, results in a potentiation of the excitability of the B-cell which derives from an increase in input resistance across the B-cell soma membrane. Pressure microapplication of gamma-aminobutyric acid (GABA) (12.5 microM) on the terminal branches of the Hermissenda Type B photoreceptors results in the fast (less than 1 s) activation of an inward Cl- conductance, characterized by a decrease in neuronal membrane resistance and an accompanying hyperpolarization (3-6 mV) of the B-cell. A slower effect of GABA, characterized by a slight depolarization (2-4 mV) and increase in resistance was observed approximately 2 min after GABA application. Following bath application of the Cl- channel blocker picrotoxin (100 microM), this increase in resistance was observed within 20 s of GABA application, suggesting that it was normally masked by the faster Cl- conductance. The magnitude of the resistance increase in response to GABA was enhanced when the B-cell was held at depolarized membrane potentials (-40 to -20 mV), but was eliminated if Ca2+ was removed from the extracellular bath, or if the non-specific protein kinase inhibitor H7 (100 microM) was added to the extracellular bath. In a final experiment, GABA application was paired with a transient (10 s) depolarization of the B-cell (to -20 mV).(ABSTRACT TRUNCATED AT 250 WORDS)     

Relationship between two voltage-dependent serotonin responses of molluscan neurones

A serotonin (5-HT)-induced slow inward current was reanalyzed in identified snail neurones and found to result from a decrease in a voltage-dependent K+-conductance, sensitive to [Ca2+]0 changes. 5-HT evoked in the same neurones an increase in the spike plateau known to be associated to a K+-conductance decrease. Both 5-HT responses appear to reflect the same decrease in K+-conductance.     

Modulation of ionic currents in Aplysia motor neuron B15 by serotonin, neuropeptides, and second messengers

Both 5-HT and the 9 amino acid neuropeptide SCPb modulate 3 ionic currents in B15, enhancing a voltage-dependent inward sodium current, decreasing an outward potassium current and increasing an inward rectifying potassium current. In contrast, FMRFamide decreases a voltage-dependent inward sodium current and increases an outward potassium current. We have also investigated the roles of several second-messenger systems that may be mediating the effects of these modulators. Bath application of membrane permeable analogs of cAMP enhance the voltage-dependent inward sodium current and both 5-HT and SCPb increase cAMP levels in B15, suggesting that cAMP may be mediating part of the observed effects of these transmitters on B15. Experiments with phorbol ester, a protein kinase inhibitor, and a phospholipase inhibitor suggest that the phospholipase C/protein kinase C cascade may decrease an outward potassium current. Thus, 5-HT and SCPb may activate multiple second-messenger systems to modulate 3 ionic currents in B15. Additional studies suggest that a cascade involving arachidonic acid may be involved in mediating part of the FMRFamide responses in B15. These studies are beginning to define molecular mechanisms whereby a neuron differentially modulates multiple ionic currents in response to distinct chemical messengers.     

Regulation of short-term associative memory by calcium-dependent protein kinase

Neural and behavioral correlates of an associative memory in Hermissenda were examined during induction and/or formation of the memory. Hermissenda received either light (conditioned stimulus or CS) and rotation (unconditioned stimulus or US) paired (i.e., Pavlovian conditioning), light and rotation unpaired (pseudoconditioning), or no exposure to light and rotation. Following 9 pairings in a 6 min session, conditioned animals exhibited a contraction of the foot in response to a test CS presented 2 min after the last conditioning trial, whereas pseudoconditioned and untreated animals exhibited a foot extension to the same CS. In addition, both an associative and a nonassociative reduction in light-induced locomotion was observed. To examine neural correlates of this learning within minutes of acquisition, the isolated nervous system of the Hermissenda (containing the visual and vestibular organs) was trained with stimulus conditions identical to those used for the intact animal. Prior isolation and preparation of the nervous system permitted immediate intracellular recording following the final conditioning trial. Relative to pseudoconditioned and untreated animals, the B photoreceptors in conditioned nervous systems were found to have elevated input resistance (inversely related to K+ channel conductance and positively related to excitability) and exhibited increased steady-state depolarization in response to the light CS, as well as a prolonged depolarization after the CS offset. These neural correlates of the associative memory were attenuated if the protein kinase inhibitor H7 was present in the extracellular bath during conditioning, demonstrating in the reduced preparation that antagonism of protein kinase activity blocks the induction of membrane alterations of identified neurons that correlate with memory storage.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in ionic conductances induced by cAMP in Helix neurons

Adenosine 3',5'-cyclic monophosphate (cAMP) was injected by a fast and quantitative pressure injection method into voltage-clamped identified Helix neurons. The intracellular elevation of cAMP caused an inward current which was not accompanied by a significant change in membrane conductance in a negative potential range with little activation of voltage-dependent membrane conductances. Near resting potential Na+ ions were the main carrier of the cAMP-induced inward current as measured with ion-selective microelectrodes. TTX did not affect the Na+ influx. K+ and less effective Ca2+ could substitute for Na+ in carrying the inward current. In the presence of Na+, divalent cations such as Ca2+ and Mg2+, and also La3+ exerted an inhibitory influence on the cAMP-induced inward current, and Ca2+ as measured with ion-selective microelectrodes did not contribute significantly to the current. Thus, the inward current was of a non-specific nature. Simultaneously to this cAMP action, the membrane permeability for K+ ions was decreased by cAMP. This effect became particularly obvious when K+ currents were activated by long-lasting, depolarizing voltage steps. In this situation a reduced K+ efflux following cAMP injection was observed by means of K+-selective microelectrodes located near the external membrane surface. Outward K+ currents were less reduced by cAMP if external Ca2+ was replaced by Ni2+. The nearly compensatory increase and decrease of two membrane conductances in the same neuron explained the lack of change in the cell input resistance despite the considerable depolarizing action of intracellularly elevated cAMP.     

Voltage clamp analysis of membrane currents in larval muscle fibers of Drosophila: alteration of potassium currents in Shaker mutants

The body wall muscles in Drosophila larvae are suitable for voltage clamp analysis of changes in membrane excitability caused by mutations. Both inward and outward ionic currents are present in these muscle fibers. The inward current is mediated by voltage-dependent Ca2+ channels. In Ca2+-free saline, the inward current is eliminated. The remaining outward K+ currents consist of two distinct components, an early transient IA and a delayed steady IK, which are separable by differences in the rate and voltage dependence of activation and inactivation. The steady-state and kinetic properties of the activation and inactivation processes of these two currents are analyzed. The results provide a basis for quantitative analysis of altered membrane currents in behavioral mutants of Drosophila. Previous studies indicate that mutations in the Shaker (Sh) locus alter excitability in both nerve and muscle in Drosophila. Our results support the idea that the channels mediating IA are molecularly distinct from those mediating IK. All Sh mutations studied specifically affect IA without changing the properties of the calcium current and IK. In certain alleles (ShKS133, Sh102, and ShM) IA is eliminated, permitting detailed studies of IK in isolation of IA. Studies of the alleles that do not eliminate IA provide additional information of the channels. In one such allele, Sh5, voltage dependence of IA activation is shifted to more positive potentials. This is accompanied by a less pronounced shift in the voltage dependence of inactivation. These results suggest that Sh5 mutation affects the voltage-sensitive mechanism of both activation and inactivation processes and that these two processes are not controlled by independent parts of the channel. Furthermore, the differential effects of these alleles on different excitable membranes imply that other genes take part in the control of IA. The effects of Sh5 on muscle depend on developmental stage. In larval muscle, Sh5 reduces the amplitude of IA because of the shift in the current-voltage (I-V) relation. In contrast, in adult Sh5 muscles, IA is reported to be normal in amplitude but shows abnormally rapid inactivation (Salkoff, L., and R. Wyman (1981) Nature 293: 228-230). A different allele, ShrK0120, causes a clear defect in nerve excitability, but analysis of IA in ShrK0120 larval muscle reveals I-V relations, inactivation, and recovery from inactivation similar to those seen in normal fibers. We suggest a possible mechanism of combinations of multiple interacting genes participating in the control of potassium channels to account for the presence of a variety of potassium channels in different excitable membranes.     

Voltage- and ligand-activated inwardly rectifying currents in dorsal raphe neurons in vitro

Intracellular recordings were made from neurons in rat dorsal raphe in the slice preparation maintained at 37 degrees C. The single-electrode voltage-clamp method was used to measure membrane currents at potentials more negative than rest (-60 mV). Three types of inward rectification were observed: 2 in the absence of any drugs and the third induced by 5-HT 1 and GABA-B receptor agonists. In the absence of any drugs, an inward current activated over 1-2 sec when the membrane potential was stepped to potentials more negative than -70 mV. This current was blocked by cesium (2 mM) and resembles IQ or IH. A second inward current (IIR) occurred at membrane potentials near the potassium equilibrium potential (EK). This inward current activated within the settling time of the clamp and was abolished by both barium (10-100 microM) and cesium (2 mM). 5-HT 1 agonists activated a potassium conductance that hyperpolarized the cells at rest. This potassium conductance was about 2 nS at -60 mV and increased linearly with membrane hyperpolarization to about 4 nS at -120 mV. Baclofen activated a potassium conductance identical in amplitude and voltage dependence to that induced by 5-HT 1 agonists. Both the baclofen- and 5-HT-induced currents were nearly abolished in animals pretreated with pertussis toxin. The results indicate that a common potassium conductance is increased by 5-HT acting on 5-HT 1 receptors and baclofen acting on GABA-B receptors. This potassium conductance rectifies inwardly and is distinct from the Q-current. The ligand-activated potassium conductance also differs from the other form of inward rectification (IIR) in its voltage dependence and sensitivity to pertussis toxin.     

The effects of serotonin on N-methyl-D-aspartate and synaptically evoked depolarizations in rat neocortical neurons

The effects of serotonin (5-HT) on neuronal responses to the excitatory amino acid agonist N-methyl-D-aspartate (NMDA) were examined in neocortical slices of the Fischer rat using current-clamp and single-electrode voltage-clamp techniques. Layer V neocortical neurons responded to application of NMDA by depolarization with no change or an apparent increase in input resistance. Following perfusion with 10(-5) M 5-HT, the response of these neurons to NMDA was significantly increased in both amplitude and duration, whereas neuronal responses to quisqualic acid and acetylcholine were not altered by 5-HT. Furthermore, the enhanced response to NMDA in 5-HT was long-lasting, and could not be reversed during the course of the experiment. Resting membrane potential and the postspike train afterhyperpolarization were not significantly altered by 5-HT, although the input resistance was decreased by 5-HT. Excitatory postsynaptic potentials (EPSPs) were usually not affected or reversibly decreased by 5-HT. However, in a few cells exhibiting a complex voltage-dependent EPSP, 5-HT produced a long-lasting enhancement in the amplitude of the EPSP. Under voltage-clamp conditions, with Na+- and K+-channels blocked, 5-HT enhanced the inward current stimulated by application of NMDA. It is suggested that 5-HT selectively enhances the voltage- and Ca2+-dependent NMDA response.     

Intrinsic light responses of retinal ganglion cells projecting to the circadian system

In mammals, light entrainment of the circadian clock, located in the suprachiasmatic nuclei (SCN), requires retinal input. Traditional rod and cone photoreceptors, however, are not required. Instead, the SCN-projecting retinal ganglion cells (RGCs) function as autonomous photoreceptors and exhibit light responses independent of rod- and cone-driven input. Using whole-cell patch-clamp recording techniques, we have investigated the morphological and electrophysiological properties of this unique class of RGCs. Although SCN-projecting RGCs resemble Type III cells in form, they display strikingly different physiological properties from these neurons. First, in response to the injection of a sustained depolarizing current, SCN-projecting cells fired in a transient fashion, in contrast to most RGCs which fired robust trains of action potentials. Second, in response to light, SCN-projecting RGCs exhibited an intensity-dependent transient depolarization in the absence of rod and cone input. This depolarization reached a peak within 5 s and generated increased spiking activity before decaying to a plateau. Voltage-clamp recordings were used to characterize the light-activated conductance which generated this depolarization. In response to varying light intensities, SCN-projecting RGCs exhibited a graded transient inward current which peaked within 5 s and decayed to a plateau. The voltage dependence of the light-activated current was obtained by subtracting currents elicited by a voltage ramp before and during illumination. The light-activated current displayed both inward and outward rectification and was largely unaffected by substitution of extracellular Na+ with choline. In both respects, the intrinsic light-activated current observed in SCN-projecting RGCs resembles currents carried by ion channels of the transient receptor potential (trp) family, which are known to mediate the light response of invertebrate photoreceptors.     

Adenosine modulates a voltage-dependent chloride conductance in cultured hippocampal neurons

Whole cell currents were recorded in cultured rat hippocampal neurons using the patch-clamp method. When the cells were held near the resting membrane potential (-60 mV) the application of adenosine (1 microM) or the adenosine analogues 2CA (100 nM) and R-PIA (40 nM) induced a steady-state inward current. This response was unchanged when extra- and intracellular media were used, in which Na+ and K+ were substituted by impermeable ions in equimolar concentrations. In contrast the current was affected by lowering the extracellular Cl- concentration and thus Cl- was considered to be the ionic carrier. Additionally an almost complete block of the current was observed after applications of DIDS (50 microM), a putative Cl- channel blocker. The modulated current was voltage-dependent and was slowly activated by hyperpolarizing voltage steps. The adenosine action was theophylline- and pertussis toxin-sensitive indicating that the modulatory effect is mediated via an A1 receptor coupled to a G protein of the Gi or Go class.     

Serotonin receptors expressed in Xenopus oocytes by mRNA from brain mediate a closing of K+ membrane channels

Membrane currents evoked by serotonin (5-HT) were studied in Xenopus oocytes injected with rat brain mRNA. Intracellular EGTA was used to abolish the Ca2(+)-dependent oscillatory Cl- current to 5-HT, revealing an underlying smooth inward current. This was associated with a decreased membrane conductance, was antagonized by Ba2+ and Zn2+ (but not TEA), and probably arises through a closing of K+ channels. Half-maximal responses were obtained with 30 nM 5-HT, while 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT) was ineffective. Furthermore, methysergide, mianserin and lysergic acid antagonized the K(+)-closing response to 5-HT, consistent with it being mediated through 5-HT1C receptors. The largest K(+)-closing responses were induced by a size fraction of mRNA which also induced a large K+ conductance, suggesting that the response requires expression of both receptors and K+ channels. The K(+)-closing response induced in the oocyte resembles the M- and S-type currents described in, respectively, mammalian and invertebrate neurons.     

Relation between metabolism and the function of ion channels in the nerve cell membrane

The effect of varying intracellular 3.5-cAMP level on steady-state and voltage-dependent transmembrane ionic currents has been studied on both vertebrate and invertebrate nerve cells. A change in 3.5-cAMP level was achieved either by its direct introduction into the cell or by stimulation or inhibition of the activity of different enzymes of the cyclase system. Intracellular cAMP increase was found to activate a steady-state transmembrane current whose early component is related mainly to the increase of sodium and calcium conductance of the membrane, and a late one to increase of potassium conductance (possibly, activated by Ca2+ influx). The decrease of intracellular cAMP concentration (by intracellular dialysis) results in reduction of the potential-activated inward calcium current, whereas its increase restores the current. Restoration of calcium current can be achieved by activation of cellular adenylate cyclase, inhibition of phosphodiesterase or by direct intracellular introduction of the catalytic subunit of cAMP-dependent protein kinase. The evidence is presented that the observed regulatory effects are mediated through cAMP-dependent phosphorylation of proteins of corresponding ionic channels. The increase of intracellular Ca2+ level closely cooperates with this regulatory metabolic system by activation of a number of its enzymatic processes.     

Pacemaker potentials of serotonergic dorsal raphe neurons: Contribution of a low-threshold Ca2+ conductance

Neurons of the dorsal raphe nucleus exhibit intrinsic pacemaker potentials (gradual interspike depolarizing ramps) enabling them to sustain spontaneous rhythmic activity in the absence of synaptic interactions. A depolarizing prepotential (PP) has been observed in these cells, which appears to trigger the spike toward the end of the pacemaker cycle. The purposes of this study, carried out in the rat dorsal raphe nucleus brain slice preparation, were to (1) determine the ionic nature of the PP, (2) investigate its time- and voltage-dependent properties, and (3) investigate the possible modulation of the underlying conductance by the α₁-agonist phenylephrine and by serotonin (5-HT), agents that modify dorsal raphe pacemaker activity. During intracellular recording under current clamp, PPs were completely and reversibly blocked by divalent cations indicating that Ca²⁺ carries a significant portion of the current causing the PPs. Ni²⁺ specifically inhibited the PP with no effect on high-threshold (?40 mV) Ca²⁺ spikes or the Ca²⁺ activated K⁺ conductance in these neurons. Activation threshold for the PP was found to be approximately ?60 mV. Priming by hyperpolarization allowed removal of inactivation (de-inactivation) of the PP in a time- and voltage-dependent manner, with maximal PPs accompanying hyperpolarizing pulses to ?90 mV and the de-inactivation beginning to occur between ?65 and ?75 mV. Single-electrode voltage-clamp experiments demonstrated a region of negative-slope conductance between ?60 and ?50 mV, which corresponds to the range of PP activation. The results of this study are consistent with a lowthreshold Ca²⁺ conductance underlying the PP whose role is to enable the membrane potential to rebound to action potential threshold at the end of the pacemaker cycle; neither phenylephrine nor 5-HT directly affected this inward current.     

Activation of neurokinin receptors modulates K and Cl channel activity in cultured astrocytes from rat cortex

Short application of the neurokinin receptor agonist substance P (SP) leads to a biphasic depolarization of astrocytes cultured from rat cortex. The rapid and transient depolarizing event lasted few seconds, the slow one several minutes. In some cells, only the slow depolarizing component was observed. During the slow depolarizing event, the sensitivity of the membrane potential for a change in the K+ gradient decreased, indicating a decrease in the relative K+ permeability of the membrane. The rapid SP-induced depolarization could be reversed, when the membrane potential was depolarized to about 0 mV by elevation of the extracellular K+ concentration, indicating a reversal potential close to the Cl- equilibrium potential. When the membrane was clamped close to the resting membrane potential using the whole-cell patch-clamp technique, SP induced a biphasic inward current with a similar time course as the SP-induced membrane depolarization. Evaluating current-to-voltage curves indicated a conductance decrease during the slow inward current with a reversal potential of the SP-dependent current close to the K+ equilibrium potential. The mean open time of single K+ channels, measured in the cell-attached configuration of the patch-clamp technique, decreased after application of SP. In contrast, the mean open time of single Cl- channels increased. We conclude that activation of neurokinin receptors in astrocytes modulates the activity of K+ and Cl- channels, leading to a complex depolarization of the membrane potential.     

Serotonin acting via cyclic AMP enhances both the hyperpolarizing and depolarizing phases of bursting pacemaker activity in the Aplysia neuron R15

Bath application of 5-HT, at concentrations below 10 microM, enhances the amplitude of the interburst hyperpolarization in the Aplysia bursting pacemaker neuron R15. It is known that 5-HT acts via cyclic AMP to produce this effect by increasing the inwardly rectifying potassium current (IR). Here, we report that further elevating the concentration of 5-HT produces an enhancement of the depolarizing phase of the burst cycle that can eventually lead to tonic spiking activity. Voltage-clamp studies reveal that high concentrations of 5-HT continue to increase IR and, in addition enhance a voltage-gated inward current active near the action potential threshold. Pharmacological treatments and ion substitution experiments demonstrate that the inward current increased by high concentrations of 5-HT is a subthreshold calcium current (ICa). The 5-HT-induced increase in ICa is mimicked by bath application of the adenylate cyclase activator forskolin or injection of 8-bromo-cyclic AMP and is potentiated by the phosphodiesterase inhibitor isobutylmethylxanthine. It is concluded that 5-HT, acting via the second messenger cyclic AMP, can increase both potassium and calcium currents in neuron R15. It is also shown that the 5-HT-induced increase in these 2 opposing voltage-gated currents not only produces complex changes in bursting activity, but also dramatically alters R15's response to inhibitory and excitatory stimuli.