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P R Benjamin

Publications and source records attributed to P R Benjamin.

At least 55 records · Page 3Linked to original sources

Training in a novel environment improves the appetitive learning performance of the snail, Lymnaea stagnalis.

The effect of novelty, an environmental background variable affecting feeding and appetitive learning performance, was examined in a behavioral study of the pond snail Lymnaea stagnalis. Transfer of snails into a novel aquatic environment (clean water) evoked exploratory behavior which manifested itself in an increased number of spontaneous rasping movements of the mouth over the second to fifth minute after exposure to the novel environment. The intensity of this behavior was much weaker in a familiar environment (used water from the home tank). Similarly, sucrose-induced feeding rates were highest when the snails were given the sucrose stimulus in a novel environment. The effectiveness of appetitive conditioning using tactile stimulus paired with food (Kemenes & Benjamin, 1989a) improved when the snails were subjected to conditioning in a novel environment. Satiety, an internal variable, suppressed the stimulating effects of the novel environment on the spontaneous, unconditioned, and conditioned feeding alike. After training in the novel environment, the conditioned response was retained for up to 12 days and thus provided a robust behavioral paradigm for the extension of the analysis to the neurophysiological mechanisms of factors affecting appetitive learning in molluscs.

Animals↗

Modulatory role for the serotonergic cerebral giant cells in the feeding system of the snail, Lymnaea. I. Fine wire recording in the intact animal and pharmacology.

1. The role of the paired serotonergic cerebral giant cells (CGCs) in the feeding system of Lymnaea was examined by electrophysiological and pharmacological techniques. 2. The firing characteristics of the CGCs were recorded by fine wires attached to their cell bodies in freely moving intact snails (in vivo recording) and their "physiological" rates of firing determined during feeding and other behaviors. 3. The mean CGC firing rates recorded in vivo varied between 1 and 20 spikes/min but never reached the average rates seen in the isolated CNS (60-120 spikes/min). Maximum rates of firing were seen during bouts of radula biting/rasping movements characteristic of the consummatory phase of feeding (15 +/- 1.69 spikes/min, mean +/- SE, range 7-20 spikes/min), with lower rates seen during locomotion (6.7 +/- 0.75 spikes/min; range 5-9 spikes/min. The cells were rarely active when the animal was quiescent (1.45 +/- 0.91 spikes/min; range 0-2 spikes/min). 4. In vivo recorded CGC firing was phase locked to the feeding movements of the animal, with spikes occurring just before the opening of the mouth, during the protraction phase of the feeding cycle. 5. Evoking firing rates on the CGCs in the isolated preparation similar to those seen in vivo during rasping movements (7-20 spikes/min) did not elicit a fictive feeding pattern in an inactive preparation. Neither did bath application of 10(-9) M serotonin (5-HT; the transmitter of the CGCs). 6. To allow the modulatory role of the CGCs to be examined during patterned activity, the fictive feeding pattern was evoked in the isolated preparation by injecting depolarizing current into a modulatory neuron, the slow oscillator (SO). 7. The tonic firing activity of the CGCs was accurately maintained by current injection in the isolated preparation at rates equivalent to that occurring during feeding, locomotion, and quiescence in the intact snail. This was possible where the CGCs became silent after 1-2 h. Only when the CGCs activity was maintained at a rate (approximately 15 spikes/min) similar to that occurring during rasping, was the SO able to drive a full, high-frequency fictive feeding pattern (15-20 cycles/min). At lower rates of CGC firing, the SO-driven rhythm was either of lower frequency or no rhythm occurred at all (CGCs silent). 8. In many isolated preparations (80%) the CGCs remained active, and it was difficult to maintain specific levels of tonic activity by current injection.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

Modulatory role for the serotonergic cerebral giant cells in the feeding system of the snail, Lymnaea. II. Photoinactivation.

1. Photoinactivation of dye-filled neurons was used to examine the modulatory role of the paired cerebral giant cells (CGCs) in the Lymnaea feeding system. 2. Both CGCs were filled with fluorescent dyes. Lucifer yellow was used for "soma" kills and injected via intracellular microelectrodes. CGC axons were retrogradely filled with 5 (6)-carboxyfluorescein (5-CF), through the cut ends of the ventro- and lateral buccal nerves, for "axonal" kills. 3. Irradiation of the CGC soma with a blue laser light (0.5 MW/m2) led to a loss of their recorded membrane potentials and the synaptic responses with their postsynaptic cells (feeding motor neurons). CGC coupling and axonal fluorescence were lost after axonal irradiation. 4. The tonic firing rate of CGC axon spikes in peripheral nerve roots following bilateral soma kills was reduced to approximately 15% of preirradiation levels (n = 2; from 52.5 +/- 3.75 spikes/min to 8.2 +/- 0.95 spikes/min; mean +/- SE) but spike activity was not completely eliminated. 5. The fictive feeding rhythm was evoked by depolarizing a modulatory neuron, the slow oscillator (SO), before and after laser irradiation. Thirty minutes after both the CGCs were irradiated (n = 8), the frequency of the SO-driven feeding rhythm was reduced. Mean fictive feeding rates were reduced from 8.3 to 4.5 cycles/min for soma kills (n = 3) and from 16.2 to 9.6 cycles/min for axonal kills (n = 5; P < 0.05). 6. The results suggest that the CGCs play a modulatory role in controlling the frequency of oscillation of the feeding central pattern generator (CPG) in Lymnaea. The SO could still drive a full fictive feeding rhythm after irradiation but at a reduced rate. At least in the soma kills, the residual spike activity retained in the distal branches of the CGCs appeared sufficient to allow the SO to drive this slow rhythm.

Animals↗

Genomic organization of the FMRFamide gene in Lymnaea: multiple exons encoding novel neuropeptides.

Based on the sequencing of genomic and cDNA clones, we were able to determine that the FMRFamide gene consists of five exons covering at least 20 kb and predict the presence of further novel peptides. The exons are alternatively spliced: exon I (hydrophobic leader sequence) to exon II (tetrapeptides) and exon I to exons III (heptapeptides), IV, and V. A cDNA clone encoding the heptapeptides is described and has also been shown to encode further novel peptides SKPYMRFamide, HDYMRFamide, and SSFPRYamide. Analysis of the right internal parietal nerve using mass spectrometry showed that the novel peptide SKPYMRFamide was cleaved from the precursor. This peptide excites neurons, suggesting a physiological function in the CNS.

Amino Acid Sequence↗

Processing of the FMRFamide precursor protein in the snail Lymnaea stagnalis: characterization and neuronal localization of a novel peptide, 'SEEPLY'.

In the pulmonate snail Lymnaea stagnalis, FMRFamide-like neuropeptides are encoded by a multi-exon genomic locus which is subject to regulation at the level of mRNA splicing. We aim to understand the post-translational processing of one resulting protein precursor encoding the tetrapeptide FMRFamide and a number of other putative peptides, and determine the distribution of the final peptide products in the central nervous system (CNS) and periphery of Lymnaea. We focused on two previously unknown peptide sequences predicted by molecular cloning to be encoded in the tetrapeptide protein precursor consecutively, separated by the tetrabasic cleavage site RKRR. Here we report the isolation and structural characterization of a novel non-FMRFamide-like peptide, the 22 amino acid peptide SEQPDVDDYLRDVVLQSEEPLY. The novel peptide is colocalized with FMRFamide in the CNS in a number of identified neuronal systems and their peripheral motor targets, as determined by in situ hybridization and immunocytochemistry. Its detection in heart excitatory motoneurons and in nerve fibres of the heart indicated that the novel peptide may play a role, together with FMRFamide, in heart regulation in the snail. The second predicted peptide, STEAGGQSEEMTHRTA (16 amino acids), was at very low abundance in the CNS and was only occasionally detected. Our current findings, suggestive of a distinct pattern of post-translational processing, allowed the reassessment of a previously proposed hypothesis that the two equivalent sequences in the Aplysia FMRFamide gene constitute a molluscan homologue of vertebrate corticotrophin releasing factor-like peptides.

Amino Acid Sequence↗

A molecularly defined cardiorespiratory interneuron expressing SDPFLRFamide/GDPFLRFamide in the snail Lymnaea: monosynaptic connections and pharmacology.

1. Neuron-specific expression of alternately spliced exons of the gene encoding the Phe-Met-Arg-Phe-NH2 (FMRFamide) family of neuropeptides and the role of encoded peptides in synaptic transmission were examined in an identified cardiorespiratory interneuron, the visceral white interneuron (VWI), in the snail Lymnaea. 2. In situ hybridization using exon-specific probes showed VWI cytoplasmic expression of the exon encoding the Lymnaea heptapeptides Gly-Asp-Pro-Phe-Leu-Arg-Phe-NH2 (GDPFLRF amide) Ser-Asp-Pro-Phe-Leu-Arg-Phe-NH2 (SDPFLRF amide) but not the exon encoding the tetrapeptides FMRFamide and Phe-Leu-Arg-Phe-NH2 (FLRFamide). 3. The absence of the tetrapeptides (FMRFamide and FLRFamide) in the VWI was indirectly confirmed by the lack of immunoreactivity to a specific antibody raised against the sequence Leu-Tyr. This sequence is present in the Lymnaea tetrapeptide precursor, but not the heptapeptide precursor. 4. The VWI has monosynaptic connections with many identifiable neurons in the CNS. These were excitatory on three clusters of identified neurons [B group (Bgp), E group (Egp), and F group (Fgp)], inhibitory on another cluster [A group (Agp)] or biphasic (excitation followed by inhibition) on a single giant neuron [right pedal dorsal 1 (RPeD1)]. 5. The role of GDPFLRFamide/SDPFLRFamide as putative neurotransmitters was examined by comparing neuronally evoked postsynaptic responses with the effects of focal peptide application. 6. The heptapeptides closely mimicked the inhibitory responses (threshold pressure pipette concentration 10(-9) M) on the Agp cells and RPeD1, including an increase in membrane conductance. FMRFamide was 1 order of magnitude less potent. GDPFLRFamide/SDPFLRFamide, applied either alone or in "cocktails" (combinations of GDPFLRFamide, SDPFLRFamide, FMRFamide, and FLRFamide), did not reproduce the excitatory effect of the VWI on the Bgp, Egp, and Fgp cells. These peptides, applied either together or separately, inhibited the cells. 7. FMRFamide or FLRFamide, but not GDPFLRFamide or SDPFLRFamide, could reproduce the initial depolarizing component of the biphasic response on RPeD1. This only occurred at concentrations of > or = 10(-4) M (10(-3) M was necessary to get spikes on RPeD1) and may not be physiologically significant. 8. We conclude that at least one so far unidentified co-transmitter must be present in the VWI to account for its full range of synaptic responses.

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A cholinergic modulatory interneuron in the feeding system of the snail, Lymnaea.

1. Pharmacological and physiological methods were used to examine the role of acetylcholine (ACh) in modulation of the Lymnaea feeding central pattern generator (CPG) by the slow oscillator (SO) interneuron. 2. Extracts of dissected SO cell bodies inhibited spontaneous ventricular contractions of the clam Mya arenaria, indicating the presence of ACh. These effects were blocked by the specific antagonist benzoquinonium chloride (10(-7) M). 3. Isolated SO cells grown in culture synthesized ACh from tritiated choline. 4. High [K+] saline induced release of synthesized ACh from cultured SO cells into the medium. 5. The specific ACh antagonist phenyltrimethylammonium (10(-4) M) blocked both excitatory, biphasic (inhibitory-excitatory) and inhibitory monosynaptic connections from the SO to feeding CPG interneurons and motor neurons. Less specific cholinergic antagonists blocked either excitatory (hexamethonium, 10(-4) M) or both excitatory and inhibitory connections (d-tubo-curarine, 10(-4) M). 6. The synaptic responses of the SO could be mimicked by brief (20 ms) pressure-pulsed application of ACh onto the cell bodies of the postsynaptic cells in high-Mg2+ saline. In normal saline, ACh elicited bursts of spikes in the N1 cells, indicating that a fictive feeding pattern had been induced in the CPG. This mimics the main mechanism by which the SO activates the CPG, which is by exciting the N1s. 7. The frequency of SO-induced fictive feeding rhythm was reduced by bath application of hexamethonium chloride to the buccal ganglia. This reduced the amplitude of the SO-->N1 excitatory synaptic response (30% of controls) and is probably the main mechanism for the reduction in the frequency of the rhythm. 8. The evidence suggests that ACh is the main neurochemical involved in allowing the SO to initiate and control the frequency of the Lymnaea feeding CPG.

Acetylcholine↗

Mutually exclusive expression of alternatively spliced FMRFamide transcripts in identified neuronal systems of the snail Lymnaea.

The FMRFamide gene of the snail Lymnaea encodes tetrapeptides (FMRFamide/FLRFamide) and heptapeptides (GDPFLRFamide/SDPFLRFamide) on separate exons. In situ hybridization probes specific to these exons were used to map the expression of the two exons in identified neuronal systems of the CNS. Analysis of more than 200 preparations showed that cytoplasmic expression of mRNA was exclusively of one type, with individual neurons expressing either the tetrapeptide or heptapeptide exon. Of the approximately 340 neurons expressing the two exons, the majority (80%) expressed the tetrapeptide exon. The tetrapeptide exon was more widespread, occurring in neurons from all 11 ganglia of the CNS. The heptapeptide was mainly confined to two ganglia (visceral and right parietal), with a small number of cells in three other ganglia. Mapping studies combined with dye marking of identified neurons showed the presence of the tetrapeptide exon in several behaviorally important networks: heart motoneurons, whole body withdrawal response motoneurons, and probably penis motoneurons as well as giant identified neurons (LP1, RPD1). The heptapeptides were prominent in two main clusters of cells (Bgp and Fgp) together with a smaller number of tetrapeptide-expressing cells.

Amino Acid Sequence↗

Cell-specific alternative RNA splicing of an FMRFamide gene transcript in the brain.

Individual neurons synthesize different peptide neurotransmitters and neuromodulators. In general, specificity is achieved by transcriptional regulation of neuropeptide-encoding genes. In Lymnaea, the FMRFamide and GDP/SDPFLRFamide neuropeptides are encoded by separate exons. Here we provide evidence that the two exons are part of the same gene and that in neurons expressing the gene the two exons are spliced onto a common upstream exon encoding a hydrophobic leader sequence. In addition, in situ hybridization data show that there is mutually exclusive cytoplasmic expression of each of the neuropeptide-encoding exons. Thus, differential neuropeptide synthesis is likely to be regulated by an alternative splicing mechanism. The cellular specificity of these splicing events is remarkable and suggests that cell-specific alternative splicing may be of major importance in establishing neuronal diversity in this system.

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Dopamine-immunoreactive neurones in the central nervous system of the pond snail Lymnaea stagnalis.

The distribution of dopamine and dopamine-immunoreactive neurones was studied in the central nervous system of the snail Lymnaea stagnalis. The results from immunocytochemical labelling were compared with those from the application of the glyoxylic acid fluorescence method and 6-hydroxydopamine-induced pigment labelling. Comparisons were also made between the number of dopamine immunoreactive neurones and the dopamine content of the ganglia, measured by high-performance liquid chromatography. Dopamine immunocytochemistry proved to be superior to the other two histochemical techniques in terms of specificity and sensitivity. The 6-hydroxydopamine-induced pigment labelling failed to prove a useful tool for the in vivo identification of all dopamine-containing neurones. The distribution and number of dopamine-immunoreactive neurones and levels of biochemically measured dopamine in specific ganglia showed a close correspondence. By using the results of the dopamine immunocytochemistry and glyoxylic acid technique, a detailed map of dopamine-containing neurones was constructed. Dopamine-containing inter- and intra-ganglionic axon tracts were also demonstrated. The mapping of dopamine-containing neurones will facilitate further neurophysiological analysis of dopaminergic neural mechanisms in Lymnaea.

Animals↗

The peptide FMRFamide activates a divalent cation-conducting channel in heart muscle cells of the snail Lymnaea stagnalis.

1. Isolated Lymnaea stagnalis heart ventricle cells contain cation-conducting channels with properties characteristic of Ca2+ channels. These channels, which carry inward Na+ currents in the absence of Ca2+, are activated by the molluscan cardioactive peptides FMRFamide and FLRFamide, and are blocked by Co2+ ions. 2. FMRFamide also activated inward Ba2+ currents at the cell's resting potential. These currents, which were also blocked by Co2+ ions, reversed at a membrane potential of +70 mV. 3. Both sodium and barium currents were initiated when the peptides were applied to the cell outside of the patch pipette indicating that a secondary messenger is likely to be involved in the FMRFamide response.

Animals↗

The whole-body withdrawal response of Lymnaea stagnalis. I. Identification of central motoneurones and muscles.

Two muscle systems mediated the whole-body withdrawal response of Lymnaea stagnalis: the columellar muscle (CM) and the dorsal longitudinal muscle (DLM). The CM was innervated by the columellar nerves and contracted longitudinally to shorten the ventral head-foot complex and to pull the shell forward and down over the body. The DLM was innervated by the superior and inferior cervical nerves and the left and right parietal nerves. During whole-body withdrawal, the DLM contracted synchronously with the CM and shortened the dorsal head-foot longitudinally. The CM and the DLM were innervated by a network of motoneurones. The somata of these cells were located in seven ganglia of the central nervous system (CNS), but were especially concentrated in the bilaterally symmetrical A clusters of the cerebral ganglia. The CM was innervated by cells in the cerebral and pedal ganglia and the DLM by cells in the cerebral, pedal, pleural and left parietal ganglia. Individual motoneurones innervated large, but discrete, areas of muscle, which often overlapped with those innervated by other motoneurones. Motoneuronal action potentials evoked one-for-one non-facilitating excitatory junction potentials within muscle fibres. No all-or-nothing action potentials were recorded in the CM or DLM, and they did not appear to be innervated by inhibitory motoneurones. The whole network of motoneurones was electrotonically coupled, with most cells on one side of the CNS strongly coupled to each other but weakly coupled to cells on the contralateral side of the CNS. This electrotonic coupling between motoneurones is probably important in producing synchronous contraction of the CM and DLM when the animal retracts its head-foot complex during whole-body withdrawal.

Animals↗

The whole-body withdrawal response of Lymnaea stagnalis. II. Activation of central motoneurones and muscles by sensory input.

The role of centrally located motoneurones in producing the whole-body withdrawal response of Lymnaea stagnalis (L.) was investigated. The motoneurones innervating the muscles used during whole-body withdrawal, the columellar muscle (CM) and the dorsal longitudinal muscle (DLM) were cells with a high resting potential (-60 to -70 mV) and thus a high threshold for spike initiation. In both semi-intact and isolated brain preparations these motoneurones showed very little spontaneous spike activity. When spontaneous firing was seen it could be correlated with the occurrence of two types of spontaneous excitatory postsynaptic potential (EPSP). One was a unitary EPSP that occasionally caused the initiation of single action potentials. The second was a larger-amplitude, long-duration (presumably compound) EPSP that caused the motoneurones to fire a burst of high-frequency action potentials. This second type of EPSP activity was associated with spontaneous longitudinal contractions of the body in semi-intact preparations. Tactile stimulation of the skin of Lymnaea evoked EPSPs in the CM and DLM motoneurones and in some other identified cells. These EPSPs summated and usually caused the motoneurone to fire action potentials, thus activating the withdrawal response muscles and causing longitudinal contraction of the semi-intact animal. Stimulating different areas of the body wall demonstrated that there was considerable sensory convergence on the side of the body ipsilateral to stimulation, but less on the contralateral side. Photic (light off) stimulation of the skin of Lymnaea also initiated EPSPs in CM and DLM motoneurones and in some other identified cells in the central nervous system (CNS). Cutting central nerves demonstrated that the reception of this sensory input was mediated by dermal photoreceptors distributed throughout the epidermis. The activation of the CM and DLM motoneurones by sensory input of the modalities that normally cause the whole-body withdrawal of the intact animal demonstrates that these motoneurones have the appropriate electrophysiological properties for the role of mediating whole-body withdrawal.

Animals↗

Neuropeptides Gly-Asp-Pro-Phe-Leu-Arg-Phe-amide (GDPFLRFamide) and Ser-Asp-Pro-Phe-Leu-Arg-Phe-amide (SDPFLRFamide) are encoded by an exon 3' to Phe-Met-Arg-Phe-NH2 (FMRFamide) in the snail Lymnaea stagnalis.

Biochemical analysis has shown the pond snail Lymnaea stagnalis to contain 2 main classes of Phe-Met-Arg-Phe-NH2 (FMRFamide)-like neuropeptides: the tetrapeptides FMRFamide and Phe-Leu-Arg-Phe-NH2 (FLRFamide), and the heptapeptides Gly-Asp-Pro-Phe-Leu-Arg-Phe-NH2 (GDP-FLRFamide) and Ser-Asp-Pro-Phe-Leu-Arg-Phe-NH2 (SDPFFRFamide). By genomic mapping and DNA sequencing, we show here that the GDP/SDPFLRFamide coding region lies 3' to the FMRFamide coding region. The absence of an initiating start methionine and the presence of good-concensus 3' and 5' splice sites suggests that the GDP/SDPFLRFamide coding region makes up 1 exon of a larger gene. In addition to 7 copies of GDPFLRFamide and 6 copies of SDPFLRFamide, the exon encoding the heptapeptides also encodes 3 novel peptides, Glu-Phe-Phe-Pro-Leu-NH2 (EFFPLamide), Ser-Asp-Pro-Tyr-Leu-Phe-Arg-NH2 (SDPYLFRamide), and Ser-Asp-Pro-Phe-Phe-Arg-Phe-NH2 (SDPFFRFamide). In contrast to the tetrapeptide FMRFamide precursor protein, the GDP/SDPFLRFamide peptides are encoded contiguously, being separated only by single basic amino acids.

Amino Acid Sequence↗

Pharmacology of the myogenic heart of the pond snail Lymnaea stagnalis.

1. We have used pharmacologic, immunologic, and biochemical techniques to examine the role of neurochemicals in modulating the myogenic heart of the snail, Lymnaea. 2. 5-HT [high-pressure liquid chromatography (HPLC) and immunocytochemistry], dopamine (HPLC), FMRFamide-related peptides (radioimmunoassay and immunocytochemistry) and substance P-related peptides (immunocytochemistry) were shown to be localized within heart tissue. 3. The pharmacologic actions of these substances on the auricle from an isolated heart preparation were examined together with other putative modulators, acetylcholine (ACh), small cardioactive peptides A and B (SCPA and SCPB), [Arg]8vasotocin (AVT), and Lymnaea native FMRFamide-related peptides [Phe-Met-Arg-Phe-NH2 (FMRFamide), Ser-Asp-Pro-Phe-Leu-Arg-Phe-NH2 (SDPFLRFamide) and Gly-Asp-Pro-Phe-Leu-Arg-Phe-NH2 (GDPFLRFamide)]. 4. The response to each substance could be distinguished by different effect on beat rate, amplitude, and diastolic tonus, as well as by the duration of responses to standard 1-min applications. ACh was inhibitory at low concentrations (threshold less than 10(-10) M) but excitatory at high concentrations (10(-6) M). AVT was alone in producing no dose-dependent response. At high concentrations (10(-4) M), AVT caused a massive tonic contraction and cessation of auricle beat. All other substances examined were excitatory. 5. Antagonists to 5-HT (cinanserin), dopamine (ergonovine), and ACh (alpha-bungarotoxin) were identified. 6. ACh, 5-HT, dopamine, and FMRFamide-related peptides all acted on the auricle at low concentrations, and the rapid onset and short duration of their excitatory effects (ACh inhibitory at low concentrations) suggested that they may have roles as neurotransmitters. SCPA and SCPB were also potent (threshold less than 10(-10) M) but produced long-duration responses suggesting a modulatory or hormonal role.

Animals↗

Regulation of heartbeat in Lymnaea by motoneurons containing FMRFamide-like peptides.

1. Intracellular stimulation of single neurons in the Lymnaea CNS was carried out to identify heart motoneurons. 2. Two of the identified motoneurons, the E heart excitor (Ehe) cells, were shown to contain Phe-Met-Arg-Phe-NH2 (FMRFamide)-like peptides by immunocytochemical staining of dye-marked neurons and by radioimmunoassay (RIA) applied to extracts of single dissected cells. 3. Bursts of spikes in the Ehe cells increased heart rate, beat amplitude and muscle tonus. This response was mimicked by perfusion of exogenous FMRFamide at low concentration (10(-6) to 5 x 10(-8) M) through the interior of the intact heart. 4. Application of selective antagonists to 5-hydroxytryptamine (5-HT) and dopamine failed to block Ehe cardiac effects. 5. Detailed evidence that the Ehe cells were heart motoneurons was obtained. 1) Anatomic mapping using the dye Lucifer yellow showed Ehe cells had peripheral projections restricted mainly to the intestinal nerve, the only nerve known to innervate the heart. 2) Perfusion of the CNS with a saline containing Co2+ blocked central chemical synapses but did not affect activity of Ehe cells on the heart. 3) Simultaneous intracellular recordings from Ehe cells and auricle muscle fibers showed unitary excitatory junction potentials following with constant latency from spikes in Ehe cells. 6. The present study elucidates the role of FMRFamide in cardioregulation and provides the first evidence that it acts as an excitatory neurotransmitter on the snail heart.

Animals↗

Excitation and inhibition of the heart of the snail, Lymnaea, by non-FMRFamidergic motoneurons.

1. The present paper extends the model of neuronal control of the Lymnaea heart by the use of intracellular recording techniques to identify further types of cardioactive neurons in the CNS that, like the previously described E heart excitor (Ehe) cells, influence the myogenic heartbeat. 2. Four new types of neuron that act on the heart are described. These are excitatory Hhe and She cells (H and S heart excitors) and the inhibitory Khi cell (K heart inhibitor). The fourth class, tonus pericardium excitor (Tpe), modulates the heart by action on pericardial tissue. 3. Pharmacologic, electrophysiological, and anatomic evidence is presented that shows that these cells are motoneurons, innervating heart muscle fibers directly: blocking central chemical synapses failed to prevent the actions of the neurons on the heart; simultaneous intracellular recordings showed unitary EJPs in heart muscle after 1:1 and with constant delay from evoked neuronal action potentials; intracellular injection of the dye Lucifer yellow showed all cells had axonal branches entering the intestinal nerve (which innervates the heart). 4. The use of selective antagonists to 5-hydroxytryptamine (5-HT) (cinanserin), dopamine (ergonovine), and acetylcholine (alpha-bungarotoxin) provided evidence that the actions of She and Hhe cells are mediated by 5-HT, whereas those of the Khi cell are mediated by acetylcholine. 5. A cyclically active network of three interneuronal inputs acting on the heart motoneurons is described. 6. One of these, input 3, is responsible for periodic excitation of the heart via its effects on the Hhe cells.

Acetylcholine↗