Decrease In Membrane Conductance Research Articles

Glutamate mediates a slow synaptic response in hippocampal slice cultures

Glutamate (GLU) mediates its 'fast' excitatory transmitter action in the brain by directly gating cation-selective ion channels ('ionotropic' receptors). However, GLU can also activate another type of receptor, coupled to phospholipase C ('metabotropic' receptor). In hippocampal cells, stimulation of this metabotropic receptor by GLU, or by a racemic mixture of (1S-3R and 1R-3S) 1-aminocyclopentyl-1,3-dicarboxylate (ACPD), induces a slower excitation mediated by inhibition of K+ currents. We have assessed whether this slow form of metabotropic receptor excitation can contribute to the effects of synaptically released GLU in hippocampal slice cultures, by recording the responses of CA3 pyramidal cells to afferent mossy fibre stimulation. When the fast ionotropic response was blocked pharmacologically, mossy fibre stimulation produced a slow depolarizing postsynaptic potential associated with a decrease in membrane conductance, a depression of the slow after-hyperpolarization following a train of action potentials, and reduced accommodation during the action potential train. Under voltage-clamp, mossy fibre stimulation produced a slow voltage-dependent inward current which resembled that produced by application of exogenous ACPD or quisqualate (QUIS), and which was occluded by these metabotropic agonists. We therefore suggest that synaptically released GLU can induce two types of postsynaptic responses: a fast excitation through activation of ionotropic receptors and a slower excitation associated with inhibition of K+ conductances through activation of metabotropic receptors. This is analogous to the dual action of acetylcholine on ionotropic (nicotinic) and metabotropic (muscarinic) receptors.

Receptors of the serotonin 1C subtype expressed from cloned DNA mediate the closing of K+ membrane channels encoded by brain mRNA.

The modulation of K+ channels by serotonin (5-HT) receptors was studied by coinjecting Xenopus oocytes with mRNA transcribed in vitro from a cloned 5-HT 1C subtype (5-HT1C) receptor gene, together with size-fractionated mRNA isolated from rat cerebral cortex that expresses K+ channels. After intracellular loading with EGTA to block Ca2(+)-dependent chloride currents, these oocytes responded to 5-HT with an inward current associated with a decrease in membrane conductance. Membrane current responses were small or absent in oocytes injected with either mRNA alone. We conclude that 5-HT1C receptors are able to cause the closing of a class of K+ channels expressed by cortex mRNA in a Ca2(+)-independent manner. The coupling between the receptors and channels appears to be mediated by the inositol phospholipid second messenger pathway, since activation of this pathway by application of serum evoked a similar closing current.

Short‐term action of insulin on Aplysia neurons: Generation of a possible novel modulator of ion channels

In mollusks as in other animals, peptides can act as hormones, growth factors, and neurotransmitters. The presence of insulin in vertebrate brain as well as its actions on nerve cells led us to examine the electrophysiological effects of the mammalian hormone on Aplysia neurons. Application of insulin extracellularly causes hyperpolarization of L14 and L10, identified neurons of the abdominal ganglion. This hyperpolarization is associated with a decreased membrane conductance that reverses at -35 mV. We also injected inositol phosphate glycan (IPG) into the identified neurons. This complex sugar, which was purified from rat liver and which is a putative second messenger for insulin in nonneural vertebrate cells (Saltiel and Cuatrecasas, 1986; Saltiel, Osterman, and Darnell, 1988), causes hyperpolarization with decreased membrane conductance in L14 and L10 similar to the effects of insulin. Furthermore, exposure of isolated ganglia to insulin results in the generation of IPG with a compensating decrease in its glycosyl-phosphatidylinositol precursor. We suggest that, in addition to its other roles, insulin may function as a neuropeptide transmitter using IPG as a second messenger.

Mechanism of Cl- sensitivity in internal ion receptors of the leech; an inward current gated off by Cl- in the nephridial nerve cells

The nephridial nerve cells of the leech, Hirudo medicinalis, 34 sensory cells, each associated with one nephridium, are sensitive to changes in extracellular Cl- concentration, an important factor in ion homeostasis. Using single-electrode current- and voltage clamp and ion substitution techniques, the specificity and mechanism of Cl- sensitivity of the nephridial nerve cell was studied in isolated preparations. Increase of the normally low external Cl- concentration leads to immediate and sustained hyperpolarization, decrease of the frequency of bursts and decrease of membrane conductance. The response is halogen specific: Cl- can be replaced by Br-, but not by organic mono- or divalent anions or inorganic divalent anions. At physiological Cl- concentrations (36 mM extracellular Cl-), the nephridial nerve cell has a high resting conductance for Cl- and the membrane potential is governed by Cl-. In high extracellular Cl- concentrations (110-130 mM), membrane conductance is low, most likely due to the gating off of Cl- channels. Under these conditions, membrane potential is dominated by the K+ distribution and the nephridial nerve cell hyperpolarizes towards EK.

Glutamate receptors of rod bipolar cells are linked to a cyclic GMP cascade via a G-protein

Whole-cell patch-clamp recordings were obtained from light-responsive on-bipolar cells in retinal slices of the dogfish. Inclusion of the G-protein activator, GTP gamma S, in the intracellular patch solution mimicked the action of glutamate, inducing an increase in net outward current (interpreted as a decrease in inward current), a decrease in membrane conductance and block of light responses. Cyclic GMP (cGMP) in the patch pipette increased inward current and membrane conductance, and blocked light responses. Cyclic AMP had no effect. IBMX, a phosphodiesterase inhibitor, produced the same effect as cGMP, suggesting the presence of a cGMP phosphodiesterase in rod bipolar cells. These results indicate that the glutamate receptors of on-bipolar cells are coupled via a G-protein to regulate intracellular cGMP, which, in turn, results in the opening of sub-synaptic membrane channels. The similarity to phototransduction is striking, and the proposed scheme would account for the high gain in transmission of rod signals to on-bipolar cells.

Activation of protein kinase C by phorbol ester induces downregulation of the Na+/K+-ATPase in oocytes ofXenopus laevis

Full-grown prophase-arrested oocytes of Xenopus laevis were treated with 50 nM phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C, or with 50 nM 4 alpha-phorbol 12,13-didecanoate (4 alpha PDD) that does not activate protein kinase C. The effect on membrane currents and capacitance, inulin uptake and ouabain binding, and on membrane morphology were analyzed. (i) During application of PMA, current generated by the Na+/K+ pump decreases; in addition, Cl- and K+ channels become inhibited. This general decrease in membrane conductance reaches steady state after about 60 min. 4 alpha PDD was ineffective. (ii) Ouabain binding experiments demonstrate that PMA (K1/2 = 7 nM), but not 4 alpha PPD, induces a reduction of the number of pump molecules in the surface membrane. Permeabilization of oocytes by digitonin plus 0.02% SDS renders all binding sites present prior to PMA treatment again accessible for ouabain. The KD value for ouabain binding is not influenced. 4 alpha PDD was ineffective. (iii) Exposure of oocytes to PMA reduces membrane capacitance and stimulates uptake of inulin suggesting an increase in endocytosis. Electron micrographs show that PMA reduces the number and length of microvilli, leading finally to a smooth membrane surface with a reduced surface area. From these results we conclude that stimulation of protein kinase C leads to downregulation of the sodium pump. A major portion of this inhibition is brought about by reduction in area of surface membrane with a concomitant internalization of pump molecules. In addition to this mode of downregulation, a direct effect of stimulation of protein kinase C on the pump molecule cannot be excluded.

GABA B receptors expressed in Xenopus oocytes by guinea pig cerebral mRNA are functionally coupled with Ca 2+-dependent Cl − channels and with K + channels, through GTP-binding proteins

Transmembrane currents induced by (−)-baclofen (BAC), a specific agonist of the γ-aminobutyric acid-B 9GABA B) receptor, in Xenopus oocytes injected with guinea pig cerebral mRNA were electrophysiologically and pharmacologically characterized under a voltage-clamp condition. The oocytes injected with mRNA acquired responsiveness to BAC and showed two types of currents at a holding potential of −50 mV. One was the slow and smooth inward current which had a short lateny and associated with a decrease in membrane conductance, and its amplitude was decreased by hyperpolarization and increased by depolarization. The other was the large fast oscillatory inward current with a long-latency, which was decreased in amplitude by depolarization and reversed at −26 mV. Both currents were not blocked by bicuculline but were depressed by 2-hydroxysaclofen (2-OH-SAC), though the smooth current was less sensitive to 2-OH-SAC; about 40% blockade at the 2-OH-SAC concentration capable of abolishing the oscillatory current. The smooth current was depressed by Ba 2+. The intracellular injection of EGTA into oocytes abolished the oscillatory current but did not affect the smooth current. The oscillatory current was time-dependently attenuated and almost abolished by intracellularly injected pertussis toxin (PTX), while the smooth current was not depressed by this toxin even when the oscillatory current was nearly abolished. The intracellular injection of GTP-γ-S into oocytes attenuated both oscillatory and smooth currents. These results suggest the possibility that GABA B receptors expressed in Xenopus oocytes by cerebral mRNA are functionally coupled with two signal transduciton systems, one is the opening of Ca 2+-dependent cl − channels mediated by PTX_sensitive GTP-binding protein(s) and the other is the closure of K + channels through PTX-insensitive GTP-binding protein(s).

Inhibition of a cAMP-dependent Ca-activated K conductance by forskolin prolongs Ca action potential duration in lamprey sensory neurons

Intracellular recordings from primary mechanosensory neurons (dorsal cells) of the lamprey spinal cord were made to test the membrane effects of forskolin, an activator of adenylate cyclase in these cells. At a concentration of 50 μM, forskolin was found to have a pronounced broadening effect on calcium action potentials (Ca APs) produced in the presence of voltage-activated K channel blockers (TEA, 3,4-DAP). Forskolin had no effect on passive membrane properties of the cells, such as resting potential or input resistance. Nor did it affect delayed rectification or Na APs and thus appeared not to block voltage-activated K channels. Forskolin's effect was evident only when a significant Ca component to the AP was present. It appeared not to increase the conductance of the Ca channel since its action was accompanied by a decrease in membrane conductance during the Ca AP, indicating instead an inhibition of a repolarizing Ca-activated conductance, other than a Ca-activated Cl conductance. The prolongation of Ca APs by forskolin, barium or the neurotransmitter GABA were all correlated in voltage-clamp with a decrease in outward current. Under the conductions used here, the major outward conductance in dorsal cells is a Ca-activated K conductance(gK(Ca))28, and it is concluded that the most probable mode of action for forskolin is via a cyclic AMP-mediated inhibition of this conductance. GABA has also been shown to prolong Ca APs in lamprey dorsal cells by inhibiting a repolarizinggK(Ca)28. Thus, the action of this transmitter may be mediated by an increase in intracellular cyclic AMP.

Variation in the pattern of [Ca2+]i change induced by acetylcholine in cultured hippocampal neurons

Acetylcholine (ACh) caused various patterns of change in the intracellular Ca2+ concentration ([Ca2+]i) in cultured rat hippocampal neurons. We studied the underlying mechanisms of the [Ca2+]i changes with simultaneous recording of [Ca2+]i and membrane potential/current. In most cases, [Ca2+]i rise was accompanied by a membrane depolarization. The [Ca2+]i change was significantly reduced when the membrane was voltage clamped, which implies that most of the [Ca2+]i rise results from the Ca2+ influx through the voltage-gated Ca2+ channel activated by the membrane depolarization. The membrane depolarizations were classified into two types, one associated with membrane conductance decrease and the other associated with membrane conductance increase. The former results from potassium conductance ((gK+) decrease, and the latter may result from the activation of a Na(+)-permeable channel. However, [Ca2+]i elevation was also observed in some neurons showing membrane hyperpolarization in response to ACh. This seems to show that ACh liberates Ca2+ from the intracellular Ca2+ store, resulting in the activation of a calcium-dependent K+ channel (KCa). The variations of ACh response in the hippocampal neurons seem to result from a variety of muscarinic acetylcholine receptors and various species of ion channels governed by those receptors.

Chara buckellii, a Euryhaline Charophyte from an Unusual Saline Environment

The time course of turgor regulation of the euryhaline giant-celled alga, Chara buckellii, is presented. Isolated intermodal cells were challenged by increasing or decreasing the external osmotic pressure by 150 milliosmoles per kilogram with all ions in the media or by dilution, respectively. Regulation following hypotonic stress was complete within 48 hours whereas regulation following hypertonic stress required between 96 and 144 hours. The change in internal osmotic pressure could be entirely accounted for by changes in vacuolar KCl in response to hypotonic stress, but this ion pair only accounted for 45% of the change in response to hypertonic stress. The membrane potential of C. buckellii is normally hyperpolarized with respect to the equilibrium potential for K(+) (E(K)). The membrane depolarized to a level close to E(K) in response to hypotonic treatment and this was accompanied by a transient increase in membrane conductance. In response to hypertonic stress, the membrane hyperpolarized transiently, then repolarized to a level close to the control. This was accompanied by a temporary decrease in membrane conductance. The data are discussed with respect to the ecological significance of the time course and ion transport mechanisms during turgor regulation.

Voltage- and transmitter-gated currents in isolated rod bipolar cells of rat retina

1. Bipolar cells were isolated from adult rat retinas after enzymatic and mechanical treatment. The cells could be unequivocally identified from their morphology because of high retention of their axon and dendritic processes after isolation. 2. Protein kinase C (PKC) immunoreactivity performed on sections of the rat retina labeled rod bipolar cells and a few amacrine cells. Virtually all bipolar cells in the dissociates expressed PKC immunoreactivity and were, therefore, rod bipolar cells. 3. Rod bipolar cells were examined with the tight-seal whole-cell and excised-patch recording techniques. Resting potentials of the isolated cells recorded under current-clamp conditions showed a broad unimodal distribution around -37 mV. 4. Membrane depolarization from a holding potential of -90 mV resulted in an outward current. A fast sodium inward current was not observed. Membrane hyperpolarization from a holding potential of -40 mV activated an inwardly rectifying current. 5. gamma-Aminobutyric acid (GABA) and glycine, the putative retinal neurotransmitters that mediate the bipolar cells' receptive field surround in vivo, activated chloride conductances in almost all isolated bipolar cells. GABA- and glycine-evoked currents were both desensitizing and could be antagonized by the classical blockers bicuculline, picrotoxin, and strychnine, respectively. 6. Pressure application of the drugs from fine microcapillaries to various parts of the isolated cells suggests a high GABA sensitivity at the axonal endings compared with either the somatic or dendritic region. A similar distribution was not found for glycine. On the contrary, glycine-induced single-channel events with main conductances of 52 and 34 pS were recorded from membrane patches excised from the cells' somata. 7. Conductances induced by glutamate and several excitatory amino acid agonists were observed in a number of the cells. Application of the glutamate agonist 2-amino-4-phosphonobutyric acid (APB) induced an inward current at negative holding potentials associated with the opening of ion channels. In only 5 of 93 cells, APB closed ion channels, leading to a decrease in membrane conductance.

The light-suppressible K+ conductance and evaluation of internal messenger candidates in the molluscan extraocular photoreceptor

A photoreceptor potential produced by a decrease in membrane conductance was not thought to occur in any invertebrate photoreceptors. However, we have found that the molluscan extraocular photoreceptor, A-P-1 responds to light with a depolarizing receptor potential due to a decrease in K+ conductance, so that the photoresponse associated with a decrease in membrane conductance is not unique to the vertebrate photoreceptor. The properties that the light-suppressible K+ conductance is time- and voltage-dependent are explained by comparison with those of the single channel conductance obtained in patch-clamp of both vertebrate and invertebrate photoreceptors. The noise analysis of the light-induced current suggest that this macroscopic light-suppressible conductance consists of channels. It is concluded that the light-suppressible K+ conductance is mediated by hydrolysis of cGMP which reduces internal cGMP, in agreement with the cGMP hypothesis of vertebrate phototransduction and that the hydrolysis may be modified by IP3.

Lymphatic Innervation

Bovine mesenteric lymphatic vessels have nerves in their walls which in response to field stimulation cause an increase in frequency of spontaneous lymphatic contractions and this could be blocked by alpha-antagonists. When vessels were loaded with [3H]-noradrenaline, 3H-efflux was increased in response to field stimulation and this was potentiated by alpha 2-antagonists and depressed by alpha 2-agonists. Electrical activity in these vessels consisted of a single action potential which preceded each contraction. Mean resting potential was -61 mV +/- 5.7 (SD). Stimulation of postsynaptic alpha-receptors caused a depolarization accompanied by a decrease in membrane conductance while beta-receptor stimulation had the opposite effect. Lymphatic noradrenergic nerves appear to have a role in the living animal since stimulation of the sympathetic chain in anaesthetized sheep increased popliteal efferent lymph flow and this could be blocked by alpha-adrenergic blockers.

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

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

Potassium currents in submucous neurones of guinea‐pig caecum and their synaptic modification.

1. Intracellular recordings were made from submucous neurones of the guinea-pig caecum. In most experiments, membrane currents were measured using a single-electrode voltage clamp. 2. A potassium current dependent on calcium influx occurred at rest (approximately equal to 200 pA at -60 mV). The amplitude of the current was increased up to 1 nA at -35 mV and decreased to zero at -100 mV; when fully activated the current did not show any inactivation. An inward calcium current, of 15-25 pA in amplitude near -60 mV and insensitive to omega-conotoxin (0.5 microM), probably activated the potassium current. 3. Step depolarizations from potentials negative to -80 mV evoked a transient (less than or equal to 200 ms at -40 mV) potassium current which was blocked by 4-aminopyridine (1-3 mM). Hyperpolarizing commands to potentials negative to -87 mV evoked an inwardly rectifying potassium current which was selectively blocked by caesium (1-2 mM). The residual cell current between -100 and -40 mV in calcium-free solution containing tetraethylammonium (20 mM), caesium (2 mM) and 4-amino-pyridine (3 mM) conformed to constant field assumptions. This current was called a background potassium current. 4. Decrease in membrane conductance during the slow excitatory postsynaptic current (EPSC) was due predominantly (greater than or equal to 90%) to a reduction in the calcium-activated potassium current at -35 mV, but due almost exclusively to a reduction in the background potassium current at potentials more negative than -100 mV. The relative contribution of the two currents to the slow EPSC was entirely dependent on the relative contribution of the currents to the membrane conductance at given potentials. 5. The transient potassium current was unaffected or slightly enhanced during the slow EPSC. The inwardly rectifying potassium current was unaffected during the slow EPSC. 6. Three tachykinins (substance P, substance K and neurokinin B; 3-800 nM), forskolin (1-30 microM), 8-bromoadenosine 3':5'-cyclic monophosphate (8-bromo cyclic AMP; 1-3 mM), 3-isobutyl-1-methylxanthine (0.3-1 mM) mimicked the conductance changes during the slow EPSC in a concentration-dependent manner. 7. It is concluded that the slow excitatory synaptic potential in the submucous plexus, presumably mediated by peptidergic transmitters, results from an inactivation of two distinct potassium currents, at least one of which is controlled by intracellular calcium ions.

Spontaneous activity mediated by NMDA receptors in immature rat entorhinal cortex in vitro

Intracellular recordings were made from cells in layer II of entorhinal cortex slices from 9 to 13-day-old rats. The majority of these cells showed pronounced spontaneous synaptic activity which could summate into large depolarizing events and give rise to bursts of spikes. These events could be associated with an apparent decrease in membrane conductance. They were reduced in amplitude by hyperpolarization of the cell and tended to increase on depolarization. Perfusion with 2-amino-5-phosphonovalerate abolished the spontaneous activity. Thus, in layer II of the entorhinal cortex at this stage of development there seems to be a functional enhancement of ongoing synaptic activity mediated via activation of N- methyl- d-aspartate receptors.

ENFLURANE-INDUCED BURST DISCHARGE OF HIPPOCAMPAL CA1 NEURONES IS BLOCKED BY THE NMDA RECEPTOR ANTAGONIST APV

The basis for the hyperexcitability and seizure activity associated with enflurane anaesthesia was investigated using extracellular and intracellular recording in rat hippocampal brain slices. Enflurane produced seizure-like burst discharges in CA1 pyramidal neurones, accompanied by depressed field potential amplitudes and a reduced threshold for synaptically evoked population spikes. However, threshold for action potentials evoked by intracellular current injection did not change, nor did action potential amplitude, duration or spike frequency accommodation in single neurones. Enflurane 2.0 vol% hyperpolarized CA1 neurones (3.1 (SD 1.3)mV), decreased membrane conductance (12 (6)% below control), and depressed EPSP amplitudes (34% of control) (P less than 0.01). Enflurane appeared to enhance both intrinsic and synaptically mediated inhibitory potentials. The N-methyl-D-aspartate (NMDA) receptor antagonist amino-phosphonovalerate (APV) 5-20 mumol litre-1 completely blocked seizure-like burst discharge of CA1 neurones in the presence of enflurane, and the enflurane-induced reduction of population spike threshold; it did not alter anaesthetic depression of EPSP amplitude. Thus enflurane-induced burst discharge of CA1 neurones appeared to involve an enhancement of excitatory synaptic transmission rather than depression of intrinsic or synaptic inhibition.

Characterization of the UV-induced electrophysiological response in smooth muscle

The effect of ultraviolet (UV) light on taenia caeci smooth muscle of guinea-pig was studied. This stimulus induced a decrease in membrane conductance, hyperpolarization of the smooth muscle cells, a decrease in spike activity and a diminished level of cGMP and enhancement of lipid peroxidation. The hyperpolarization evoked by UV light was attenuated in the presence of sodium nitrite, reaching a maximum at 0.5 mM sodium nitrite. The level of cGMP was enhanced in the presence of nitrite and lipid peroxidation was not changed. Under these conditions lipid peroxidation was not changed during UV irradiation either. The UV-induced hyperpolarization was abolished in the absence of extracellular calcium, inhibited in the presence of 8-Br-cGMP and enhanced by an increase in the extracellular calcium concentration. These results are consistent with the supposition that UV-A irradiation blocks calcium influx via a cGMP-dependent mechanism.

Endogenous histamine excites neurones in the guinea-pig superior cervical ganglion in vitro.

1. Intracellular recordings were obtained from neurones in the guinea-pig superior cervical ganglion (SCG) in vitro to study the electrophysiological effects of endogenously released histamine. 2. Guinea-pigs were actively sensitized to the specific antigen, ovalbumin. SCG removed from these animals rapidly released a significant proportion of their endogenous histamine stores into the extracellular space upon exposure to the sensitizing antigen. Several observations indicated that the released histamine was derived from ganglionic mast cells. 3. The electrophysiological effects produced by antigen challenge in a neurone mimicked qualitatively and quantitatively those effects produced by exogenously applied histamine in the same neurone. Under current clamp the membrane effects of antigen and histamine included a transient depolarization, an increase in input resistance and transient blockade of a long-duration component of the spike after-hyperpolarization. In voltage clamp histamine and antigen produced an inward current and decreased membrane conductance. 4. Histamine H1, but not H2 or H3 receptor antagonists prevented the membrane depolarization to both histamine and antigen treatments. 5. These convergent biochemical, physiological and pharmacological data demonstrate that a sufficient quantity of endogenous histamine is released by an antigenic stimulus in SCG from sensitized guinea-pigs to affect specific electrophysiological characteristics of neurones. Histamine may thus be involved in mediating interactions between the mammalian immune system and the peripheral sympathetic nervous system.

Sequence of pedal peptide: a novel neuropeptide from the central nervous system of Aplysia

We report the identification of a novel neuropeptide from Aplysia nervous tissue. The peptide was termed Pedal peptide (Pep) because it was predominantly synthesized in the pedal ganglia. Pep was purified and sequenced from pooled extracts of pedal ganglia. The following sequence was proposed: Pro-Leu-Asp-Ser-Val-Tyr-Gly-Thr-His-Gly-Met-Ser-Gly-Phe-Ala. Enzymatic hydrolysis procedures indicated that Pep had a free carboxyl terminal. A peptide with the proposed sequence was synthesized and compared with the native peptide. Chromatographic properties of the 2 peptides under 3 different conditioned were compared and found to be identical. Electrophysiological responses to the 2 peptides were compared on an identified neuron in the abdominal ganglia and found to be qualitatively and quantitatively very similar. Both peptides produced net inward currents that were associated with a decrease in membrane conductance. The results from these 2 procedures confirmed that the proposed Pep sequence was correct. Quantitative measurements of the incorporation of 35S-methionine into Pep suggest that cell bodies that synthesize Pep were present predominantly in the pedal ganglia but should also be found in other central ganglia as well. Pep-like immunoreactive neurons are found predominantly in the pedal ganglia and less frequently in the other ganglia (Pearson and Lloyd, 1989). Quantitatively, Pep constitutes one of the predominant peptides in the nervous system of Aplysia. Pep does not appear to be a member of any other previously identified invertebrate or vertebrate peptide family.