Feeding Behavior In Aplysia Research Articles

Full Hill-type muscle model of the I1/I3 retractor muscle complex in Aplysia californica.

The coordination of complex behavior requires knowledge of both neural dynamics and the mechanics of the periphery. The feeding system of Aplysia californica is an excellent model for investigating questions in soft body systems' neuromechanics because of its experimental tractability. Prior work has attempted to elucidate the mechanical properties of the periphery by using a Hill-type muscle model to characterize the force generation capabilities of the key protractor muscle responsible for moving Aplysia's grasper anteriorly, the I2 muscle. However, the I1/I3 muscle, which is the main driver of retractions of Aplysia's grasper, has not been characterized. Because of the importance of the musculature's properties in generating functional behavior, understanding the properties of muscles like the I1/I3 complex may help to create more realistic simulations of the feeding behavior of Aplysia, which can aid in greater understanding of the neuromechanics of soft-bodied systems. To bridge this gap, in this work, the I1/I3 muscle complex was characterized using force-frequency, length-tension, and force-velocity experiments and showed that a Hill-type model can accurately predict its force-generation properties. Furthermore, the muscle's peak isometric force and stiffness were found to exceed those of the I2 muscle, and these results were analyzed in the context of prior studies on the I1/I3 complex's kinematics in vivo.

Specific Plasticity Loci and Their Synergism Mediate Operant Conditioning.

Despite numerous studies examining the mechanisms of operant conditioning (OC), the diversity of OC plasticity loci and their synergism have not been examined sufficiently. In the well-characterized feeding neural circuit of Aplysia, in vivo and in vitro appetitive OC increases neuronal excitability and electrical coupling among several neurons leading to an increase in expression of ingestive behavior. Here, we used the in vitro analog of OC to investigate whether OC reduces the excitability of a neuron, B4, whose inhibitory connections decrease expression of ingestive behavior. We found OC decreased the excitability of B4. This change appeared intrinsic to B4 because it could be replicated with an analog of OC in isolated cultures of B4 neurons. In addition to changes in B4 excitability, OC decreased the strength of B4's inhibitory connection to a key decision-making neuron, B51. The OC-induced changes were specific without affecting the excitability of another neuron critical for feeding behavior, B8, or the B4-to-B8 inhibitory connection. A conductance-based circuit model indicated that reducing the B4-to-B51 synapse, or increasing B51 excitability, mediated the OC phenotype more effectively than did decreasing B4 excitability. We combined these modifications to examine whether they could act synergistically. Combinations including B51 synergistically enhanced feeding. Taken together, these results suggest modifications of diverse loci work synergistically to mediate OC and that some neurons are well suited to work synergistically with plasticity in other loci.SIGNIFICANCE STATEMENT The ways in which synergism of diverse plasticity loci mediate the change in motor patterns in operant conditioning (OC) are poorly understood. Here, we found that OC was in part mediated by decreasing the intrinsic excitability of a critical neuron of Aplysia feeding behavior, and specifically reducing the strength of one of its inhibitory connections that targets a key decision-making neuron. A conductance-based computational model indicated that the known plasticity loci showed a surprising level of synergism to mediate the behavioral changes associated with OC. These results highlight the importance of understanding the diversity, specificity and synergy among different types of plasticity that encode memory. Also, because OC in Aplysia is mediated by dopamine (DA), the present study provides insights into specific and synergistic mechanisms of DA-mediated reinforcement of behaviors.

Computational model of the distributed representation of operant reward memory: combinatoric engagement of intrinsic and synaptic plasticity mechanisms.

Operant reward learning of feeding behavior in Aplysia increases the frequency and regularity of biting, as well as biases buccal motor patterns (BMPs) toward ingestion-like BMPs (iBMPs). The engram underlying this memory comprises cells that are part of a central pattern generating (CPG) circuit and includes increases in the intrinsic excitability of identified cells B30, B51, B63, and B65, and increases in B63–B30 and B63–B65 electrical synaptic coupling. To examine the ways in which sites of plasticity (individually and in combination) contribute to memory expression, a model of the CPG was developed. The model included conductance-based descriptions of cells CBI-2, B4, B8, B20, B30, B31, B34, B40, B51, B52, B63, B64, and B65, and their synaptic connections. The model generated patterned activity that resembled physiological BMPs, and implementation of the engram reproduced increases in frequency, regularity, and bias. Combined enhancement of B30, B63, and B65 excitabilities increased BMP frequency and regularity, but not bias toward iBMPs. Individually, B30 increased regularity and bias, B51 increased bias, B63 increased frequency, and B65 decreased all three BMP features. Combined synaptic plasticity contributed primarily to regularity, but also to frequency and bias. B63–B30 coupling contributed to regularity and bias, and B63–B65 coupling contributed to all BMP features. Each site of plasticity altered multiple BMP features simultaneously. Moreover, plasticity loci exhibited mutual dependence and synergism. These results indicate that the memory for operant reward learning emerged from the combinatoric engagement of multiple sites of plasticity.

The significance of dynamical architecture for adaptive responses to mechanical loads during rhythmic behavior.

Many behaviors require reliably generating sequences of motor activity while adapting the activity to incoming sensory information. This process has often been conceptually explained as either fully dependent on sensory input (a chain reflex) or fully independent of sensory input (an idealized central pattern generator, or CPG), although the consensus of the field is that most neural pattern generators lie somewhere between these two extremes. Many mathematical models of neural pattern generators use limit cycles to generate the sequence of behaviors, but other models, such as a heteroclinic channel (an attracting chain of saddle points), have been suggested. To explore the range of intermediate behaviors between CPGs and chain reflexes, in this paper we describe a nominal model of swallowing in Aplysia californica. Depending upon the value of a single parameter, the model can transition from a generic limit cycle regime to a heteroclinic regime (where the trajectory slows as it passes near saddle points). We then study the behavior of the system in these two regimes and compare the behavior of the models with behavior recorded in the animal in vivo and in vitro. We show that while both pattern generators can generate similar behavior, the stable heteroclinic channel can better respond to changes in sensory input induced by load, and that the response matches the changes seen when a load is added in vivo. We then show that the underlying stable heteroclinic channel architecture exhibits dramatic slowing of activity when sensory and endogenous input is reduced, and show that similar slowing with removal of proprioception is seen in vitro. Finally, we show that the distributions of burst lengths seen in vivo are better matched by the distribution expected from a system operating in the heteroclinic regime than that expected from a generic limit cycle. These observations suggest that generic limit cycle models may fail to capture key aspects of Aplysia feeding behavior, and that alternative architectures such as heteroclinic channels may provide better descriptions.

Implication of dopaminergic modulation in operant reward learning and the induction of compulsive-like feeding behavior inAplysia

Feeding in Aplysia provides an amenable model system for analyzing the neuronal substrates of motivated behavior and its adaptability by associative reward learning and neuromodulation. Among such learning processes, appetitive operant conditioning that leads to a compulsive-like expression of feeding actions is known to be associated with changes in the membrane properties and electrical coupling of essential action-initiating B63 neurons in the buccal central pattern generator (CPG). Moreover, the food-reward signal for this learning is conveyed in the esophageal nerve (En), an input nerve rich in dopamine-containing fibers. Here, to investigate whether dopamine (DA) is involved in this learning-induced plasticity, we used an in vitro analog of operant conditioning in which electrical stimulation of En substituted the contingent reinforcement of biting movements in vivo. Our data indicate that contingent En stimulation does, indeed, replicate the operant learning-induced changes in CPG output and the underlying membrane and synaptic properties of B63. Significantly, moreover, this network and cellular plasticity was blocked when the input nerve was stimulated in the presence of the DA receptor antagonist cis-flupenthixol. These results therefore suggest that En-derived dopaminergic modulation of CPG circuitry contributes to the operant reward-dependent emergence of a compulsive-like expression of Aplysia's feeding behavior.

ARPOP: an appetitive reward-based pseudo-outer-product neural fuzzy inference system inspired from the operant conditioning of feeding behavior in Aplysia.

Appetitive operant conditioning in Aplysia for feeding behavior via the electrical stimulation of the esophageal nerve contingently reinforces each spontaneous bite during the feeding process. This results in the acquisition of operant memory by the contingently reinforced animals. Analysis of the cellular and molecular mechanisms of the feeding motor circuitry revealed that activity-dependent neuronal modulation occurs at the interneurons that mediate feeding behaviors. This provides evidence that interneurons are possible loci of plasticity and constitute another mechanism for memory storage in addition to memory storage attributed to activity-dependent synaptic plasticity. In this paper, an associative ambiguity correction-based neuro-fuzzy network, called appetitive reward-based pseudo-outer-product-compositional rule of inference [ARPOP-CRI(S)], is trained based on an appetitive reward-based learning algorithm which is biologically inspired by the appetitive operant conditioning of the feeding behavior in Aplysia. A variant of the Hebbian learning rule called Hebbian concomitant learning is proposed as the building block in the neuro-fuzzy network learning algorithm. The proposed algorithm possesses the distinguishing features of the sequential learning algorithm. In addition, the proposed ARPOP-CRI(S) neuro-fuzzy system encodes fuzzy knowledge in the form of linguistic rules that satisfies the semantic criteria for low-level fuzzy model interpretability. ARPOP-CRI(S) is evaluated and compared against other modeling techniques using benchmark time-series datasets. Experimental results are encouraging and show that ARPOP-CRI(S) is a viable modeling technique for time-variant problem domains.

Feeding behavior of Aplysia: A model system for comparing cellular mechanisms of classical and operant conditioning

Feeding behavior of Aplysia provides an excellent model system for analyzing and comparing mechanisms underlying appetitive classical conditioning and reward operant conditioning. Behavioral protocols have been developed for both forms of associative learning, both of which increase the occurrence of biting following training. Because the neural circuitry that mediates the behavior is well characterized and amenable to detailed cellular analyses, substantial progress has been made toward a comparative analysis of the cellular mechanisms underlying these two forms of associative learning. Both forms of associative learning use the same reinforcement pathway (the esophageal nerve, En) and the same reinforcement transmitter (dopamine, DA). In addition, at least one cellular locus of plasticity (cell B51) is modified by both forms of associative learning. However, the two forms of associative learning have opposite effects on B51. Classical conditioning decreases the excitability of B51, whereas operant conditioning increases the excitability of B51. Thus, the approach of using two forms of associative learning to modify a single behavior, which is mediated by an analytically tractable neural circuit, is revealing similarities and differences in the mechanisms that underlie classical and operant conditioning.

Cycle-to-cycle variability as an optimal behavioral strategy

Aplysia feeding behavior is highly variable from cycle to cycle. In some cycles, when the variability causes a mismatch between the animal's movements and the requirements of the feeding task, the variability makes the behavior unsuccessful. We propose that the behavior is variable nevertheless because the variability serves a higher-order functional purpose. When the animal is faced with a new and only imperfectly known feeding task in each cycle, the variability implements a trial-and-error search through the space of possible feeding movements. Over many cycles, this may be the animal's optimal strategy in an uncertain and changing feeding environment.

Classical and operant conditioning differentially modify the intrinsic properties of an identified neuron

A long-standing debate in neuroscience is whether classical and operant conditioning are mechanistically similar or distinct. The feeding behavior of Aplysia provides a model system suitable for addressing this question. Here we report that classical and operant conditioning of feeding behavior differentially modify the intrinsic excitability of neuron B51, a critical element for the expression of the feeding response, thus revealing that these two forms of associative learning differ at the cellular level.

3P212 アメフラシ摂食、吐き出し応答を誘発する海藻アミノ酸成分の探索(化学受容)

3P212 アメフラシ摂食、吐き出し応答を誘発する海藻アミノ酸成分の探索(化学受容)

A population of pedal-buccal projection neurons associated with appetitive components of Aplysia feeding behavior.

Backfills of the cerebral-buccal connective (CBC) of Aplysia californica revealed a cluster of five to seven pedal-buccal projection neurons in the anterolateral quadrant of the ventral surface of each pedal ganglion. Intra- and extracellular recordings showed that the pedal-buccal projection neurons shared common electrophysiological properties and synaptic inputs. However, they exhibited considerable heterogeneity with respect to their projection patterns. All pedal-buccal projection neurons that were tested received a slow excitatory postsynaptic potential from the ipsi- and contralateral cerebral-pedal regulator (C-PR) neuron, a cell that is thought to play a key role in the generation of a food-induced arousal state. Tests were conducted to identify potential synaptic follower neurons of the pedal-buccal projection neurons in the cerebral and buccal ganglia, but none were detected. Finally, nerve recordings revealed projections from the pedal-buccal projection neurons in the nerves associated with the buccal ganglion. In tests designed to determine the functional properties of these peripheral projections, no evidence was obtained supporting a mechanosensory or proprioceptive role and no movements were observed when they were fired. It is proposed that peripheral elements utilized in consummatory phases of Aplysia feeding may be directly influenced by a neuronal pathway that is activated during the food-induced arousal state.

Classical Conditioning of Feeding inAplysia: I. Behavioral Analysis

A training protocol was developed to classically condition feeding behavior in Aplysia californica using tactile stimulation of the lips as the conditional stimulus (CS) and food as the unconditional stimulus (US). Paired training induced a greater increase in the number of bites to the CS than unpaired training or US-only stimulation. Memory for classical conditioning was retained for at least 24 hr. The organization of the reinforcement pathway that supports classical conditioning was analyzed in additional behavioral experiments. No evidence was found for the contribution to appetitive reinforcement of US-mediating pathways originating in the lips of the animals. Bilateral lesions of the anterior branch of the esophageal nerve, which innervates parts of the foregut, however, were found to attenuate classical conditioning. Thus, it appears likely that reinforcement during appetitive classical conditioning of feeding was mediated by afferent pathways that originate in the foregut. The companion paper () describes two neurophysiological correlates of the classical conditioning.

Classical Conditioning of Feeding inAplysia: II. Neurophysiological Correlates

Feeding behavior in Aplysia californica can be classically conditioned using tactile stimulation of the lips as conditional stimulus (CS) and food as unconditional stimulus (US) [ (companion paper)]. Conditioning resulted in an increase in the number of CS-evoked bites that persisted for at least 24 hr after training. In this study, neurophysiological correlates of classical conditioning training were identified and characterized in an in vitro preparation of the cerebral and buccal ganglia. Stimulation of a lip nerve (AT(4)), which mediates mechanosensory information, resulted in a greater number of buccal motor patterns (BMPs) in ganglia isolated from animals that had received paired training than in ganglia from control animals. The majority of the evoked BMPs were classified as ingestion-like patterns. Intracellular recordings from pattern-initiating neuron B31/32 revealed that stimulation of AT(4) evoked greater excitatory input in B31/32 in preparations from animals that had received paired training than from control animals. In contrast, excitatory input to buccal neuron B4/5 in response to stimulation of AT(4) was not significantly increased by paired training. Moreover, correlates of classical conditioning were specific to stimulation of AT(4). The number of spontaneously occurring BMPs and the intrinsic properties of two buccal neurons (B4/5 and B31/32) did not differ between groups. These results suggest that appetitive classical conditioning of feeding resulted in the pairing-specific strengthening of the polysynaptic pathway between afferent fibers and pattern-initiating neurons of the buccal central pattern generator.

TheAplysia MytilusInhibitory Peptide-Related Peptides: Identification, Cloning, Processing, Distribution, and Action

Neuropeptides are a ubiquitous class of signaling molecules. In our attempt to understand the generation of feeding behavior in Aplysia, we have sought to identify and fully characterize the neuropeptides operating in this system. Preliminary evidence indicated that Mytilus inhibitory peptide (MIP)-like peptides are present and operating in the circuitry that generates feeding in Aplysia. MIPs were originally isolated from the bivalve mollusc Mytilus edulis, and related peptides have been identified in other invertebrate species, but no precursor has been identified. In this study, we describe the isolation and characterization of novel Aplysia MIP-related peptides (AMRPs) and their precursor. Several AMRPs appear to have some structural and functional features similar to vertebrate opioid peptides. We use matrix-assisted laser desorption/ionization time-of-flight mass spectrometry to confirm that all 14 AMRPs predicted by the precursor are processed in isolated neurons. Northern analysis, whole-mount in situ hybridization, and immunohistochemistry are used to map the abundant expression of these peptides in the CNS and peripheral tissues such as the digestive tract, vasculature, and the reproductive organs. Physiological studies demonstrate that the rank order of the inhibitory actions of these peptides is different for three target muscles. These results underscore the importance of using a multidisciplinary approach to identifying and characterizing the actions of neuropeptides in an effort to gain understanding of their role in systems of interest. The widespread distribution of the AMRPs indicates that they may be operating in many different systems of Aplysia.

Cloning and Characterization ofAplysiaNeutral Endopeptidase, a Metallo-Endopeptidase Involved in the Extracellular Metabolism of Neuropeptides inAplysia californica

Cell surface metallo-endopeptidases play important roles in cell communication by controlling the levels of bioactive peptides around peptide receptors. To understand the relative relevance of these enzymes in the CNS, we characterized a metallo-endopeptidase in the CNS of Aplysia californica, whose peptidergic pathways are well described at the molecular, cellular, and physiological levels. The membrane-bound activity cleaved Leu-enkephalin at the Gly3-Phe4 bond with an inhibitor profile similar to that of the mammalian neutral endopeptidase (NEP). This functional homology was supported by the molecular cloning of cDNAs from the CNS, which demonstrated that the Aplysia and mammalian NEPs share all the same amino acids that are essential for the enzymatic activity. The protein is recognized both by specific anti-Aplysia NEP (apNEP) antibodies and by the [125I]-labeled NEP-specific inhibitor RB104, demonstrating that the apNEP gene codes for the RB104-binding protein. In situ hybridization experiments on sections of the ganglia of the CNS revealed that apNEP is expressed in neurons and that the mRNA is present both in the cell bodies and in neurites that travel along the neuropil and peripheral nerves. When incubated in the presence of a specific NEP inhibitor, many neurons of the buccal ganglion showed a greatly prolonged physiological response to stimulation, suggesting that NEP-like metallo-endopeptidases may play a critical role in the regulation of the feeding behavior in Aplysia. One of the putative targets of apNEP in this behavior is the small cardioactive peptide, as suggested by RP-HPLC experiments. More generally, the presence of apNEP in the CNS and periphery may indicate that it could play a major role in the modulation of synaptic transmission in Aplysia and in the metabolism of neuropeptides close to their point of release.

Serotonergic neurons differentially modulate the efficacy of two motor neurons innervating the same muscle fibers in Aplysia.

Feeding behavior in Aplysia shows substantial plasticity. An important site for the generation of this plasticity is the modulation of synaptic transmission between motor neurons and the buccal muscles that generate feeding movements. We have been studying this modulation in the anterior portion of intrinsic buccal muscle 3 (I3a), which is innervated by two excitatory motor neurons and identified serotonergic modulatory neurons, the metacerebral cells (MCCs). We have shown previously that serotonin (5-HT) applied selectively to the muscle potently modulates excitatory junction potentials (EJPs) and contractions. All the effects of 5-HT were persistent, lasting many hours after wash out. We examined whether the release of endogenous 5-HT from the MCC could produce effects similar to the application of 5-HT. Stimulation of the MCCs did produce similar short-term effects to the application of 5-HT. MCC stimulation facilitates EJPs, potentiates contractions, and decreases the latency between the onset of a motor neuron burst and the onset of the evoked contraction. The effects of MCC stimulation reached a maximum at quite low firing frequencies, which were in the range of those previously recorded during feeding behavior. The maximal effects were similar to those produced by superfusion with approximately 0.1 microM 5-HT. Although the effects of MCC stimulation on EJPs were persistent, they were less persistent than the effects of 0.1 microM 5-HT. Mechanisms that may account for differences in the persistence between released and superfused 5-HT are discussed. Thus activity in the MCCs has dramatic short-term effects on the behavioral output of motor neurons, increasing the amplitude and relaxation rate of contractions evoked by both B3 and B38 and shifting the temporal relationship between B38 bursts and evoked contractions.

Ion currents and mechanisms of modulation in the radula opener muscles of Aplysia.

Ion currents and mechanisms of modulation in the radula opener muscles of Aplysia. J. Neurophysiol. 78: 2372-2387, 1997. Numerous studies of plasticity in the feeding behavior of Aplysia have shown that substantial plasticity is due to peripheral neuromodulation of the feeding musculature. Extensive previous work focusing on the accessory radula closer (ARC) muscle has led to the realization that a major function of the modulation in that muscle may be to ensure efficient coordination between its contractions and those of its antagonist muscles. For a more complete understanding, therefore, we must study these muscles also. Here we have studied the radula opener muscles I7-I10. Using single isolated muscle fibers under voltage clamp, we have characterized ion currents gated by voltage and by the physiological contraction-inducing neurotransmitter acetylcholine (ACh) and the effects of the physiological modulators serotonin, myomodulins A and B, and FMRFamide. Our results explain significant aspects of the electrophysiological behavior of the whole opener muscles, as well as why the opener and ARC muscles behave similarly in many ways yet differently in some key respects. Opener muscles express four types of K currents: inward rectifier, A-type [IK(A)], delayed rectifier [IK(V)], and Ca2+-activated [IK(Ca)]. They also express an L-type Ca current [ICa] and a leakage current. ACh activates a positive-reversing cationic current [IACh(cat)] and a negative-reversing Cl current [IACh(Cl)]. The opener muscles differ from the ARC in that, in the openers, activation of IK(A) occurs approximately 9 mV more positive and there is much less IACh(Cl). In both muscles, IACh(cat) most likely serves to depolarize the muscle until ICa activates to supply Ca2+ for contraction, but further depolarization and spiking is opposed by coactivation of IK(A), IK(V), IK(Ca), and IACh(Cl). Thus the differences in IK(A) and IACh(Cl) may well be key factors that prevent spikes in the ARC but often allow them in the opener muscles. As in the ARC, the modulators enhance ICa and so potentiate contractions. They also activate a modulator-specific K current, which causes hyperpolarization and depression of contractions. Finally, in the opener muscles but not in the ARC, the modulators activate a depolarizing cationic current that may help phase-advance the contractions. Each modulator exerts these effects to different degrees and thus has a distinct effect on voltage and contraction size and shape. The overall effect then will depend on the specific combinations of modulators released in different behaviors. By understanding the modulation in the opener muscles, as well as in the ARC, we are now in a position to understand how the behavior of the two muscles is coordinated under a variety of circumstances.

Ganglionic distribution of inputs and outputs of C-PR, a neuron involved in the generation of a food-induced arousal state in Aplysia.

Cerebral neuron C-PR is thought to play an important role in the appetitive phase of feeding behavior of Aplysia. Here, we describe the organization of input and output pathways of C-PR. Intracellular dye fills of C-PR revealed extensive arborization of processes within the cerebral and the pedal ganglia. Numerous varicosities of varying sizes may provide points of synaptic inputs and outputs. Blocking polysynaptic transmission in the cerebral ganglion eliminated the sensory inputs to C-PR from stimuli applied to the rhinophores or tentacles, indicating that this input is probably mediated by cerebral interneurons. Identified cerebral mechanoafferent sensory neurons polysynaptically excite C-PR. Stimulation of the eyes and rhinophores with light depresses C-PR spike activity, and this effect also appears to be mediated by cerebral interneurons. C-PR has bilateral synaptic actions on numerous pedal ganglion neurons, and also has effects on cerebral neurons, including the MCC, Bn cells, CBIs and the contralateral C-PR. Although the somata of these cerebral neurons are physically close to C-PR, experiments using high divalent cation-containing solutions and cutting of various connectives indicated that the effects of C-PR on other cerebral ganglion neurons (specifically Bn cells and the MCC) are mediated by interneurons that project back to the cerebral ganglion via the pedal and pleural connectives. The indirect pathways of C-PR to other cerebral neurons may help to ensure that consummatory motor programs are not activated until the appropriate appetitive motor programs, mediated by the pedal ganglia, have begun to be expressed.

Enhancement of Ca current in the accessory radula closer muscle of Aplysia californica by neuromodulators that potentiate its contractions

A major goal of neuroscience is to identify the neural and cellular mechanisms of behavior and its plasticity. Progress toward this goal has come particularly from work with a small number of tractable model preparations. One of these is the simple neuromuscular circuit consisting of the accessory radula closer (ARC) muscle of the mollusk Aplysia californica and its innervating motor and modulatory neurons. Contraction of the ARC muscle underlies a component of Aplysia feeding behavior, and plasticity of the behavior is in large part due to modulation of the amplitude and duration of the contractions of the muscle by a variety of modulatory neurotransmitters and peptide cotransmitters, among them the small cardioactive peptides (SCPs), myomodulins (MMs), and serotonin (5-HT). We have studied single dissociated ARC muscle fibers in order to determine whether modulation of membrane ion currents in the muscle might underlie these effects. First, we confirmed that the dissociated fibers were functionally intact: just as with the whole ARC muscle, their contractions were potentiated by 5-HT and SCPB and potentiated as well as depressed by MMA, and their cAMP content was greatly elevated by 5-HT, SCPA and SCPB, and to a lesser extent by MMA and MMB. Next, using voltage-clamp techniques, we found that two ion currents present in the fibers were indeed modulated. The fibers possess a dihydropyridine-sensitive, high-threshold "L"-type Ca current. This current was enhanced by the modulators that potentiate ARC-muscle contractions--5-HT, SCPA and SCPB, and MMA and MMB--but not by buccalinA, a modulator that does not act directly on the ARC muscle. All of the potentiating modulators, as well as elevation of cAMP in the fibers by forskolin or a cAMP analog, maximally enhanced the current about twofold and mutually occluded each other's effects. Since the Ca current supplies Ca2+ necessary for contraction of the muscle, the enhancement of the current is a good candidate to be a major mechanism of the potentiation of the contractions. In the following article we report that the modulators also, to different degrees, activate a distinctive K current and thereby depress the contractions. Net potentiation or depression then depends on the balance between the relative strengths of the modulation of the two ion currents.

Characterization of the membrane ion currents of a model molluscan muscle, the accessory radula closer muscle of Aplysia california. I. Hyperpolarization-activated currents.

1. The simple neuromuscular circuit consisting of the accessory radula closer (ARC) muscle of the mollus Aplysia californica together with its innervating motor and modulatory neurons has been extensively studied as a model preparation in which it might be possible to reach an integrated understanding of the neural and cellular mechanisms of behavioral plasticity, in this case of a component of Aplysia feeding behavior. Previous work has suggested that much of the plasticity of this behavior is implemented by appropriate release of modulatory neurotransmitters and peptide cotransmitters that modulate several parameters of the contractions of the ARC muscle. However, little is as yet known about the underlying cellular mechanisms. 2. We have begun to study single, functionally intact fibers dissociated from the ARC muscle to assess to what extent the modulation of its contraction might be mediated by one candidate mechanism, modulation of its membrane ion currents. First, however, it was necessary to gain a thorough understanding of the unmodulated currents and their likely roles in normal contraction. Using voltage-clamp techniques, we have therefore identified and characterized the major currents present in the ARC muscle fibers. We describe these currents in this and the following two papers. These results constitute the first detailed description of ion currents in a molluscan muscle and lay the foundation for further study, to be presented in subsequent papers, of the roles of two currents that we have indeed found to be modulated in ways likely to contribute to the modulation of contraction. 3. In this paper we first describe the general electrophysiological characteristics of the dissociated fibers and present evidence that the fibers can be adequately space clamped. 4. The physiological operating voltage range of the nonspiking ARC muscle most likely extends from about -80 to about -25 mV. The steady-state current-voltage (I-V) relation of total membrane current rectifies inwardly in the negative and outwardly in the positive portion of this voltage range, with a plateau region of high or even negative slope resistance separating the two regions of rectification. 5. The current responsible for the inward rectification at negative voltages is a classical inwardly rectifying K current. It is activated by hyperpolarization with quasi-instantaneous kinetics; its whole I-V relation shifts along the voltage axis in a Nernstian manner with altered extracellular K+ concentration; it is blocked by low extracellular Ba2+ and Cs+, and the block is promoted by hyperpolarization.(ABSTRACT TRUNCATED AT 400 WORDS)