An identified histaminergic neuron can modulate the outputs of buccal-cerebral interneurons in Aplysia via presynaptic inhibition

We have identified 2 buccal-cerebral interneurons (BCIs), B17 and B18, that appear to be involved in the coordination of feeding behavior in Aplysia. The BCIs have their cell bodies in the buccal ganglion, but send axons to the cerebral ganglion via the cerebral-buccal connectives. The BCIs appear to make monosynaptic connections with neurons in the cerebral ganglion that modulate extrinsic muscles involved in feeding behavior. B17 and B18 are activated antiphasically during a motor program induced by stimulating the esophageal nerve and appear to "read out" different phases of the buccal program to different cells in the cerebral ganglion. B17 and B18 are not necessary, and probably not sufficient, to generate the buccal program. These BCIs, and other cells like them in the buccal ganglion, may be capable of coordinating the activity of the intrinsic muscles of the buccal mass with the activity of its extrinsic muscles, and perhaps with those of the lips, mouth, and tentacles. Identified histaminergic neuron, C2, can modulate the outputs of the BCIs onto their synaptic followers in the cerebral ganglion. Firing of C2 inhibits spiking of the BCIs, probably via cerebral-buccal interneurons. C2 also decreases the size of the EPSP that B17 and B18 evoke in cerebral neuron C4. C2 appears to do so monosynaptically, and it decreases the conductance of C4, ruling out one possible postsynaptic mechanism of action. Variance analysis of the EPSPs evoked by B18 supports the hypothesis that C2 acts presynaptically to decrease the release of transmitter. Applications of histamine to the solution bathing the neuron mimic the effect of firing C2 and reduce the size of the EPSPs B18 induces in C4. The bath-applied histamine appears to act directly on B18, since it elicits a voltage-dependent increased conductance hyperpolarization recorded in the soma of B18, and the hyperpolarization persists in a solution in which synaptic transmission has been blocked. Histamine did not produce any marked changes of the duration of a TEA-broadened somatic action potential of B18. To the extent that the soma of B18 reflects the membrane properties of its synaptic terminal region, the data suggest that histamine may produce presynaptic inhibition by hyperpolarizing the synaptic terminal region.     

Activity of an identified histaminergic neuron, and its possible role in arousal of feeding behavior in semi-intact Aplysia

The possible functions of histaminergic neuron C2 were studied in an isolated head preparation from which it was possible to obtain intracellular recordings while the buccal mass exhibited feeding-like responses. Application of food to the lips of the isolated head preparation elicited rhythmic buccal movements that appeared to be ingestion responses, since they moved seaweed into the buccal cavity and towards the esophagus, and their frequency and regularity was similar to the ingestion responses studied in a group of intact animals. The ingestion responses of the buccal mass consisted of 2 main phases of movement of the radula from a middle rest position: forward and return to rest, and backward and return to rest. The relative magnitudes and timing of these 2 phases were variable. Intracellular recordings from C2 in the isolated head revealed that C2 is silent when the buccal mass is quiescent, but that it can be excited into spike activity, either by mechanical stimulation of the perioral zone or by chemostimulation that results in rhythmic movement. C2 fires a burst of spikes in phase with each protraction-retraction cycle, and, if the movements continue, C2 fires even when the eliciting stimulus has been removed. Activity of the cell was usually preceded by fast depolarizing responses that appeared to be blocked axon spikes. The evidence suggests that C2 is part of a positive feedback loop that may help maintain the persistence of arousal of feeding behavior beyond the time that food stimuli are removed.     

Multiple roles of a histaminergic afferent neuron in the feeding behavior of Aplysia.

The cellular and circuit properties of individual identified neurons in invertebrates can be readily studied; hence it is possible to determine how the complex properties of nerve cells function in the generation of behavior. Recent studies of the cellular basis of feeding behavior in the marine mollusc Aplysia have focused on a neuron, C2, that has a variety of complex properties that determine the behavioral functions of the neuron. C2 conveys mechanosensory information from the mouth of the animal. It receives a complex pattern of inputs during feeding behavior, and generates diverse outputs that may shape behavior. It can act to filter out slow or sporadic sensory inputs, and its own outputs can be 'gated' by synaptic input. C2 uses histamine as its transmitter, and some of its synaptic outputs are modulatory and contribute to the expression of an arousal state induced by food. Other outputs shape feeding behavior directly by affecting motor neurons, as well as presynaptically inhibiting the outputs of feeding motor programs. Thus, the complex properties of this neuron may contribute to the flexibility and adaptability of feeding in Aplysia. Studies of C2 have expanded our concepts of the properties of sensory neurons.     

Modulatory synaptic actions of an identified histaminergic neuron on the serotonergic metacerebral cell of Aplysia

Possible sources of excitatory synaptic input to the serotonergic metacerebral cell (MCC) were determined by stimulating various neurons in the cerebral ganglion. Firing of the previously identified histaminergic neuron C2 was found to produce synaptic input to the MCC. The synaptic input consists of fast excitatory-inhibitory synaptic potentials on a background of a slow EPSP. The slow EPSP appears to be monosynaptic and chemically mediated since it persists in a solution of high divalent cations; broadening of the presynaptic spike enhances the EPSP; the size of the EPSP is a function of the Mg2+ and Ca2+ concentrations of the bathing solution; and the EPSP can be mimicked by application of histamine to the MCC. The slow EPSP, in addition to firing the MCC, can increase the excitability of the cell, even under conditions in which C2 is fired at a rate too slow to produce a measurable EPSP when the MCC is at rest potential. This property appears to be due to the fact that the slow EPSP results from an apparent decrease of membrane conductance so that the size of the EPSP increases markedly as the cell is depolarized, and the EPSP appears to be highly voltage-dependent so that it is small or absent close to the rest potential of the MCC. When the MCC is voltage-clamped, application of histamine to the bath results in an inward current that disappears when the MCC is hyperpolarized. The potential at which the histamine-induced current reverses or disappears is dependent on the concentration of external potassium, suggesting that, at least in part, the slow EPSP is due to a decrease of potassium conductance. The data on C2 are consistent with its being an element of the neuronal system that mediates a state of food arousal in Aplysia.     

Biochemical and morphological correlates of transmitter type in C2, an identified histaminergic neuron in Aplysia

There is compelling evidence that histamine serves as a neurotransmitter in C2, a pair of symmetrical neurons in the cerebral ganglion of Aplysia californica. These cells had previously been shown to contain high concentrations both of histamine and of its biosynthetic enzyme, histidine decarboxylase; in addition, 3H-histamine injected intrasomatically was found to move along C2's axons by fast transport. Furthermore, several actions of C2 on identified follower cells were simulated by the application of histamine. We have now characterized this identified neuron further. C2 converts 3H-histidine to histamine: 16% of the labeled precursor was converted to histamine 1 hour after intrasomatic injection. Synthesis of 3H-histamine is specific, since no conversion occurred after injection of other identified Aplysia neurons that are known to use other neurotransmitter substances. We also examined the fine structure of C2's cell body, axons, and axon terminals within the cerebral ganglion and in the nerves that carry its three peripheral branches, identified after injection of Lucifer Yellow, 3H-histamine, or horseradish peroxidase. Characteristic dense-core vesicles are present in all regions of the neuron, and are labeled after intrasomatic injection of 3H-histamine. These 100-nm vesicles together with 60-nm electron-lucent vesicles fill the varicose extensions of C2's neurites that are widely distributed within the ganglion, but only the smaller vesicles cluster at the membrane specializations presumed to be active zones that make contact with many neurons. The widespread distribution of axon terminals and varicosities is consistent with the idea that C2 is modulatory in function; 3H-histamine is taken up selectively by the cell body and axons of C2 and of several other putative histaminergic neurons in a Na+ -dependent manner. Characterization of these biochemical and morphological features of C2 adds to the large amount of information already available to make this identified cell a standard for identifying other neurons that use histamine as a transmitter.     

Neuropeptide cotransmitters released from an identified cholinergic motor neuron modulate neuromuscular efficacy in Aplysia

Intrinsic buccal muscle 5 (I5) in Aplysia is innervated by 2 motor neurons (termed B15 and B16). In addition to the classical transmitter ACh, B15 also contains the 2 neuropeptides SCPA and SCPB. In a previous study, we demonstrated that the SCPs were released from the terminals of B15 in the I5 muscle and that this release was sufficient to raise cAMP levels in I5 muscle fibers. Significant peptide release occurred only when B15 was stimulated at high frequency or at lower frequencies with a relatively long burst duration (Whim and Lloyd, 1989). In the present article, we examine the possibility that the SCPs released from B15 modulate I5 muscle contractions produced by stimulation of the second motor neuron, B16. Application of exogenous SCPs to I5 muscles increased the amplitude and relaxation rate of B16-evoked contractions. Stimulation of B15 using paradigms that have been shown previously to cause release of the SCPs resulted in a long-lasting increase in the amplitude and relaxation rate of muscle contractions evoked by B16. This modulation is unlikely to be due to the B15-induced muscle contractions themselves, because modulation of B16-evoked contraction amplitude and relaxation rate was observed when the contractions were blocked transiently by a cholinergic antagonist during B15 stimulation. Conversely, stimulation of B15 at frequencies that produce no measurable release of the SCPs did not elicit significant modulation of B16-evoked contractions. The minimum B15 stimulation frequency required to elevate muscle cAMP levels or to modulate B16-evoked contractions was found to be within the physiological range at which B15 fires during feeding. Therefore, the mechanism underlying the modulation of B16-evoked contractions by B15 is likely to involve the release of the SCPs from B15 terminals in the I5 muscle. With respect to behavior, this modulation of muscle contractions would be most likely to occur during food-induced arousal when both motor neurons fire at high frequency with brief interburst intervals.     

Sensory function and gating of histaminergic neuron C2 in Aplysia

This paper explores the possible sensory function of the identified histaminergic neuron C2. Mechanical stimulation of a narrow region around the mouth of the animal (perioral zone) elicits brief depolarizing potentials in C2. Extracellular recordings from the peripheral axons of C2 indicate that the depolarizing potentials are due to action potentials that are conveyed from the periphery but do not invade the cell body, since they fail at a region with a low safety factor within the cerebral ganglion. These blocked axonal spikes (A-spikes) function as if they were excitatory synaptic inputs to C2, since the synaptic output of C2 does not occur unless the A-spikes succeed in evoking full action potentials in the soma (or an electrically close initial segment) of C2. Furthermore, like synaptic potentials, the A-spikes exhibit temporal and spatial summation, and facilitation. C2 receives both tonic and phasic inhibitory synaptic potentials, which can decrease the summation of A-spikes and thereby alter the frequency-filtering properties of C2 or block its synaptic output. Thus, C2 appears to be an unusual proprioceptive afferent that has a high degree of integrative function and may provide critical gating that is dependent on a variety of external and internal conditions.     

Peptidergic modulation of neuronal circuitry controlling feeding in Aplysia

We examined the effects of 3 neuropeptides and the bioactive amine 5-HT on identified motoneurons (B15 and B16) and interneurons (B4, B5) involved in the control of feeding behavior in Aplysia californica. The application of egg-laying hormone (ELH), small cardioactive peptide b (SCPb), and 5-HT elicits distinct patterns of synaptically induced bursting in the neurons, while PheMetArgPheamide (FMRFamide) inhibits firing due to synaptic activity. Repetitive IPSPs recorded in B15 and B16 are induced by 5-HT and SCPb and inhibited by FMRFamide. The substances also may act directly: In solutions that block synaptic transmission SCPb excites B15, ELH excites B16, 5-HT excites B15, B16, and B4, and FMRFamide both inhibits B15 and B16 and excites B4. We suggest that the output of a buccal ganglion central pattern generator may be modulated to produce distinct patterns of motoneuron activity by these candidate transmitters. We also noted differences in the intrinsic properties of the 2 motoneurons. B15 contains SCPb immunoreactivity while B16 does not. This finding suggests that B15 may be the source for the SCPb immunoreactivity previously found at the ARC muscle and that SCPb may be acting in an autocrine mode. Also, B15 has a significantly lower resting potential than B16 and contains a large transient outward (Ia-like) current. The candidate transmitters act by exciting or inhibiting elements at every level within the hierarchically organized motor system that controls feeding. This expands the diversity of behavioral repertoires that may be elicited from a particular neural circuit.     

Dopaminergic neuron B20 generates rhythmic neuronal activity in the feeding motor circuitry ofAplysia

We have identified a buccal neuron (B20) that exhibits dopamine-like histofluorescence and that can drive a rhythmic motor program of the feeding motor circuitry of Aplysia. The cell fires vigorously during episodes of patterned buccal activity that occur spontaneously, or during buccal programs elicited by stimulation of identified cerebral command-like neurons for feeding motor programs. Preventing B20 from firing, or firing B20 at inappropriate times, can modify the program driven by the cerebral feeding command-like neuron CBI-2. When B20 is activated by means of constant depolarizing current it discharges in phasic bursts, and evokes a sustained coordinated rhythmic buccal motor program. This program incorporates numerous buccal and cerebral neurons associated with aspects of feeding responses. The B20-driven program can be reversibly blocked by the dopamine-antagonist ergonovine, suggesting that dopamine may be causally involved in the generation of the program. Although firing of B20 evokes phasic activity in cerebral command-like neurons, the presence of the cerebral ganglion is not necessary for B20 to drive the program. The data are consistent with the notion that dopaminergic neuron B20 is an element within the central pattern generator for motor programs associated with feeding.     

Characterization of glycolipids synthesized in an identified neuron of Aplysia californica

Because radioactive precursors can be injected directly into the cell body or axon of R2, a giant, identified neuron of the Aplysia abdominal ganglion, it was possible to show that glycolipid is synthesized in the cell body, inserted into membranes along with glycoprotein, and then exported into the axon within organelles that are moved by fast axonal transport. After intrasomatic injection of N-[3H]-acetyl-D-galactosamine, five major 3H-glycolipids were identified using thin layer polysilicic acid glass fiber chromatography. At least two of the lipids are negatively charged. Analysis of 32P-labeled lipid from the abdominal ganglion revealed the presence of 2-aminoethylphosphonate, indicating that these polar substances are sphingophosphonoglycolipids. The major 3H-glycolipids synthesized in R2 are similar to a family of phospholipids isolated from the skin of A. kurodai, previously characterized by Araki et al. (Araki, S., Y. Komai, and M. Satake (1980) Biochem J. 87: 503-510). Since sialic acid is absent in Aplysia as in other invertebrates, these polar glycolipids may function like gangliosides in vertebrates. The polar 3H-glycolipids are synthesized and incorporated into intracytoplasmic membranes solely in the cell body. Direct injection of the labeled sugar into the axon revealed no local synthesis or exchange of glycolipid. Moreover, there was no indication for transfer from glial cells into axoplasm. Although the incorporation of N-[3H]-acetyl-D-galactosamine into glycolipid is not affected by anisomycin, an effective inhibitor of protein synthesis, the export into the axon of membranes containing the newly synthesized lipid is completely blocked by the drug.(ABSTRACT TRUNCATED AT 250 WORDS)     

Expression of diverse neuropeptide cotransmitters by identified motor neurons in Aplysia

Neuropeptide synthesis was determined for individual identified ventral-cluster neurons in the buccal ganglia of Aplysia. Each of these cells was shown to be a motor neuron that innervates buccal muscles that generate biting and swallowing movements during feeding. Individual neurons were identified by a battery of physiological criteria and stained with intracellular injection of a vital dye, and the ganglia were incubated in 35S-methionine. Peptide synthesis was determined by measuring labeled peptides in extracts from individually dissected neuronal cell bodies analyzed by HPLC. Previously characterized peptides found to be synthesized included buccalin, FMRFamide, myomodulin, and the 2 small cardioactive peptides (SCPs). Each of these neuropeptides has been shown to modulate buccal muscle responses to motor neuron stimulation. Two other peptides were found to be synthesized in individual motor neurons. One peptide, which was consistently observed in neurons that also synthesized myomodulin, is likely to be the recently sequenced myomodulin B. The other peptide was observed in a subset of the neurons that synthesize FMRFamide. While identified motor neurons consistently synthesized the same peptide(s), neurons that innervate the same muscle often express different peptides. Neurons that synthesized the SCPs also contained SCP-like activity, as determined by snail heart bioassay. Our results indicate that every identified motor neuron synthesizes a subset of these methionine-containing peptides, and that several neurons consistently synthesize peptides that are likely to be processed from multiple precursors.     

Input-output relationships of identified buccal neurones involved in feeding control in Aplysia

A group of about 28 neurones located in the lateral portion of the caudal face of Aplysia buccal ganglion and projecting into the cerebro-buccal connective were identified by retrograde cobalt staining, and designated as L neurones. It was found that the L neurones did not establish synaptic relations with the known buccal neurones, which are mainly involved in the production of the consummatory phase of feeding, nor with several cerebral neurones tested, including the well-known serotonin giant cell. Neither did they show responses to stimulation of the nerves directed to the buccal mass. On the other hand, the L neurones showed depolarizing responses, with the possible addition of a weak, slower hyperpolarizing phase, to stimulation of the ipsi- and contralateral oesophageal nerves, which innervate the portion of the gut posterior to the buccal mass. These findings, together with several properties of the oesophageal nerve input, suggest that one function of the L cells is to transmit information about gut regions posterior to the buccal mass towards the cerebral ganglia, and that they may mediate the inhibitory influence which in Aplysia is known to be exerted upon feeding by the presence of bulk in the anterior gut. The L neurones showed synaptic responses - consisting mainly or exclusively of depolarizations - to stimulation of the cerebro-buccal connectives. Besides this, large, tonic EPSPs, which often occurred in the 'spontaneous' activity of the L neurones, were found to be generated by spikes that travelled in the cerebro-buccal connective towards the buccal ganglion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Stereotyped neuropil branching of an identified stomatogastric motor neuron

Anatomical studies of the crab stomatogastric ganglion (STG) have suggested only minimal organization within the neuropil of this structure. Here, we present evidence that, for at least one intrinsic neuron type, the ventricular dilator (VD) neuron, a highly organized and stereotyped branching structure exists within the stomatogastric neuropil. Specifically, we show the morphology of the VD neuron consists of a single primary neurite that projects from the soma into the neuropil and bifurcates into a pair of subprimary neurites, which in turn exit the neuropilar region, one entering the left and the other the right medial ventricular nerve. Nearly all secondary neurite branching of the VD neuron is from the subprimary neurites. There are approximately 22 secondary branches/neuron (range 14-28), with no significant difference between the number of secondary branches off the right vs. the left subprimary neurite, although the ratio of secondary branches between subprimaries varies (range 0.4-1.6). The fine neurites that branch from the secondary processes segregate hemispherically within the neuropil, based on the subprimary neurite of origin. Within this hemispherical organization, another level of fine neurite segregation is present, namely, the fine neurites derived from each secondary branch are restricted to discrete regions of the hemisphere with only minimal overlap with those derived from other secondary branches. Monte Carlo simulations show that this segregation differs significantly from a random distribution. The organization of branching seen in the VD neuron may play a critical role in the electrotonic and local computational organization of this neuron and sets the stage for physiological experimentation addressing these issues.     

Activity of an identified serotonergic neuron in free moving Aplysia correlates with behavioral arousal

Extracellular recordings of the metacerebral cell (MCC), a serotonergic neuron inAplysia, were obtained in free moving, undrugged animals. MCC activity was evoked by exposure to food. Arousal level was manipulated by satiating the animals or exposing them to a noxious stimulus. We found that the amount of evoked MCC activity correlated with the level of arousal of the animal.     

Time course of structural changes at identified sensory neuron synapses during long-term sensitization in Aplysia

We have used the gill- and siphon-withdrawal reflex of Aplysia californica to explore the morphological basis of the synaptic plasticity that underlies long-term sensitization. In earlier studies (Bailey and Chen, 1983, 1988a), we described 2 classes of structural changes at identified sensory neuron synapses that occur following long-term sensitization: (1) increases in the number, size, and vesicle complement of active zones and (2) an overall increase in the total number of synaptic varicosities per sensory neuron. In the present study, we have begun to examine which of these anatomical changes might be necessary for the maintenance of long-term sensitization by exploring the time course over which they occur and, in particular, their duration relative to the persistence of the memory assessed behaviorally. Toward this end we have quantitated changes in both the total number of varicosities and their active zone morphology in single HRP-labeled sensory neurons taken from long-term sensitized and control animals at different intervals (1-2 d, 1 week, and 3 weeks) following training. We have found that long-term sensitized animals examined within 48 hr after the completion of training demonstrate an increase in the total number of varicosities per sensory neuron as well as an increase in the incidence, size, and vesicle complement of their synaptic active zones compared with control animals. The increase in the number of varicosities and active zones persists unchanged for at least 1 week, and the increase in active zone number is only partially reversed at the end of 3 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)     

Proteins rapidly transported to the synapses of a single identified neuron of Aplysia californica

The cell body of R2, a giant cholinergic neuron of Aplysia californica, resides in the abdominal ganglion, whereas its synapses are on thousands of unicellular mucus glands located in the skin. Due to the great spatial separation between the site of macromolecular synthesis and the presynaptic terminals, rapid axonal transport can be used to segregate synaptic proteins from those to be used elsewhere in the cell. The proteins of R2 were labeled by incubating the abdominal ganglion in [35S]methionine for 5 hr in a chamber separated from the rest of the isolated central nervous system. After 50 hr, 28 radiolabeled proteins were reproducibly found by one- and two-dimensional polyacrylamide gel electrophoresis to be transported to the distal regions of peripheral nerves P6, P7, and P8 that innervate the parapodia and middle body wall. We are sure that R2 is the source of these proteins since radioautography of sections taken throughout the nervous system, complemented by cobalt tracings, showed that R2 is the only neuron in the abdominal ganglion with axons in these nerves. Nine of the 28 transported proteins are glycoproteins since they were also labeled after injecting R2's cell body with [3H]-L-fucose. There is evidence that the proteins and glycoproteins are destined for R2's presynaptic terminals. For example, in experiments in which the body wall and parapodium remained attached to the nerves, the proteins were transported to the skin region that contains the glands. Moreover, analyses of the distribution of the rapidly transported proteins by qualitative radioautography and by extrusion of axoplasm indicated that none are constituents of the axolemma.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cellular and synaptic morphology of a feeding motor circuit in Aplysia californica

The cellular and synaptic morphology of a component of the feeding motor circuit in Aplysia californica was examined with light and electron microscopic techniques. The circuit consists of a pair of inhibitory premotor interneurons, B4 and B5, as well as two motoneurons, B15 and B16, which innervate the accessory radula closer muscle. The neurons have wide, varicose arborizations in the buccal ganglion neuropil. All four of these neurons are cholinergic, and in addition, B15 contains immunoreactivity to sera raised against small cardioactive peptide B. Varicose processes in the accessory radula closer muscle are immunoreactive with antisera against several neuropeptides. We identified specific neuromuscular junctions by visualizing horseradish peroxidase uptake in recycled synaptic vesicles. Direct innervation of the accessory radula closer muscle by B15 and B16 is demonstrated by experiments in which horseradish peroxidase is transported from motoneuronal soma to the terminals on muscle fibers. In addition, specific synaptic contacts between B4 and B5 and each of the motoneurons are observed in the buccal ganglion neuropil. Finally, multiple contacts consistent with peptidergic, serotoninergic, and cholinergic synapses are made onto the neurons, suggesting that a variety of transmitters modulate motor output at each level of the hierarchical circuit. These results support the physiological evidence suggesting the involvement of neuropeptides as well as "classical" transmitters in the modulation of circuitry governing feeding behavior in Aplysia.     

Nitric oxide induces cGMP immunoreactivity and modulates membrane conductance in identified central neurons of Aplysia

Nitric oxide (NO) acts as an orthograde neurotransmitter in the central nervous system by stimulating guanylyl cyclase and increasing cGMP. We previously demonstrated this pathway for an identified synaptic follower neuron in the cerebral ganglion of the mollusc Aplysia californica. Here, we investigated the NO--cGMP pathway in other Aplysia central neurons using cGMP immunocytochemistry and intracellular recordings. NO-induced cGMP immunoreactivity in a few neurons in the abdominal, pleural and buccal ganglia, including identified neurons L11 and R15 in the abdominal ganglion and neuron Bng in the buccal ganglion. NO depolarized all these neurons but none of the four nonimmunoreactive neurons tested. In neuron L11, NO-induced depolarization, tonic spiking and a reduction in membrane conductance at resting potential. The NO effect was mimicked by applying the membrane permeable analogue, 8-Br-cGMP in L11. Neuron R15 was depolarized and its activity shifted from spike/bursts to tonic spiking. These NO effects were blocked by applying the guanylyl cyclase inhibitor ODQ and mimicked by 8-Br-cGMP. Neuron Bng was depolarized and produced spikes tonically. These results provide evidence that the NO--cGMP pathway is linked to membrane ionic channels in cGMP-IR neurons, and the channels may be different in specific neurons.     

Reciprocal modulation of calcium current by serotonin and dopamine in the identified Aplysia neuron R15

Voltage-clamp methods were employed to study the effects of serotonin (5-HT) and dopamine on the pharmacologically isolated calcium current in the identified Aplysia neuron R15 grown in cell culture. Neurons were obtained from juvenile animals and had not yet developed the bursting pacemaker pattern of activity characteristic of R15 in mature animals. In R15 5-HT elicits a biphasic response consisting of excitatory depolarization followed by an inhibitory hyperpolarization and dopamine elicits an inhibitory hyperpolarization. 5-HT increased the Ca2+ current without affecting its voltage dependence. The 5-HT effect persisted when Ba2+ was employed to carry current through Ca2+ channels. 5-HT did not affect the rate of Ca2+-dependent Ca2+ current inactivation other than through its effect on the magnitude of the Ca2+ current. The adenylate cyclase activator forskolin, in the presence of a phosphodiesterase inhibitor, also increased the magnitude of the Ca2+ or Ba2+ current. This result suggested that the 5-HT-induced enhancement of Ca2+ current was mediated by cAMP. Dopamine inhibited Ca2+ current when either Ca2+ or Ba2+ was employed as the current carrier. Dopamine did not affect the rate of Ca2+-dependent inactivation of Ca2+ current other than through its effect on the magnitude of the Ca2+ current. Intracellular injection of the Ca2+ chelator EGTA inhibited serotonergic modulation of the Ca2+ current but not dopaminergic modulation. These results indicated that the putative neurotransmitters 5-HT and dopamine may regulate bursting activity in mature R15 neurons through modulation of Ca2+ current.