PubMed Health⌕ Search

Biomedical subjects

V F Castellucci

Publications and source records attributed to V F Castellucci.

At least 55 records · Page 3Linked to original sources
PubMed1982

Inhibitor of adenosine 3':5'-monophosphate-dependent protein kinase blocks presynaptic facilitation in Aplysia.

Sensitization of the gill withdrawal reflex results from presynaptic facilitation at the excitatory synapses made by sensory neurons on gill motor neurons. Facilitation is accompanied by an increase in the duration of the action potential in sensory cells because of the depression of a K+ current. This results in an increasd influx of CA2+ and a greater release of transmitter from sensory neurons. There is evidence that serotonin is the facilitating transmitter and that the depression of the K+ current by serotonin mediated by cAMP-dependent protein phosphorylation. To test further the role of the cAMP-dependent protein kinase and of protein phosphorylation in sensitization, we have attempted to prevent or reverse the development of the electrophysiological correlates that accompany sensitization. We have pressure-injected sensory neurons with a specific and a stable protein inhibitor of the cAMP-dependent protein kinase both before and after the application of serotonin or the activation of the facilitator neurons. The increase in spike broadening that accompanies facilitation was prevented or diminished by injection of the inhibitor. Moreover, injection of the inhibitor could reverse fully the developed spike broadening produced by prior application of serotonin. These observations strenthen the evidence for the involvement of protein phosphorylation in presynaptic facilitation. Phosphorylation of the substrate protein evidently is quite labile and does not persist after the kinase is inhibited. Thus, the time course of short term sensitization appears to be determined by an active kinase. We think that it is likely that the mechanism for maintaining the kinase in an active form resides in the slow decay of the cAMP produced by the action of serotonin or the facilitator neurons on the sensory cells.

Animals↗
PubMed1982

Facilitatory transmitter causes a selective and prolonged increase in adenosine 3':5'-monophosphate in sensory neurons mediating the gill and siphon withdrawal reflex in Aplysia.

Sensitization of the gill and siphon withdrawal reflex in the marine mollusc, Aplysia california, is a simple form of learning Underlying this behavioral changes is a cascade of biochemical events. The first step in this cascade is postulated to be an increase in cAMP within the sensory neurons of the abdominal ganglion. We have developed a labeling protocol with 32Pi which permits us to measure the synthesis of cAMP within a single sensory neurons. Application of serotonin for 5 min was found to triple the content of [32P]cAMP in sensory neurons. The response is specific to serotonin: dopamine, a transmitter that does not produce sensitization, did not increase cAMP. Physiological stimulation of facilitator neurons also resulted in a 3.5-fold increase of cAMP in sensory neurons but not in other cells of the ganglion. We studied the time course of the increase of cAMP in sensory cells stimulated with serotonin and found that it parallels closely the time course of the short term form of presynaptic facilitation. We also have determined the effects of transmitters on the synthesis of cAMP in other identified neurons of the ganglion. The bag cells responded specifically to serotonin. R15, which has been shown to be hyperpolarized both the serotonin and by dopamine, responded to both transmitters by increased synthesis synthesis of cAMP. Thus, the dopamine- and serotonin-sensitive cyclase can be localized to both the same and different cells. Other cells did not respond to serotonin or to dopamine, indicating that a transmitter-sensitive adenylate cyclase is a specific property and is not present in all neurons.

Animals↗
PubMed1980

Presynaptic membrane potential affects transmitter release in an identified neuron in Aplysia by modulating the Ca2+ and K+ currents.

We have examined the relationships between the modulation of transmitter release and of specific ionic currents by membrane potential in the cholinergic interneuron L10 of the abdominal ganglion of Aplysia californica. The presynaptic cell body was voltage-clamped under various pharmacological conditions and transmitter release from the terminals was assayed simultaneously by recording the synaptic potentials in the postsynaptic cell. When cell L10 was voltage-clamped from a holding potential of -60 mV in the presence of tetrodotoxin, graded transmitter release was evoked by depolarizing command pulses in the membrane voltage range (-35 mV to + 10 mV) in which the Ca(2+) current was also increasing. Depolarizing the holding potential of L10 results in increased transmitter output. Two ionic mechanisms contribute to this form of plasticity. First, depolarization inactivates some K(+) channels so that depolarizing command pulses recruit a smaller K(+) current. In unclamped cells the decreased K(+) conductance causes spike-broadening and increased influx of Ca(2+) during each spike. Second, small depolarizations around resting potential (-55 mV to -35 mV) activate a steady-state Ca(2+) current that also contributes to the modulation of transmitter release, because, even with most presynaptic K(+) currents blocked pharmacologically, depolarizing the holding potential still increases transmitter release. In contrast to the steady-state Ca(2+) current, the transient inward Ca(2+) current evoked by depolarizing clamp steps is relatively unchanged from various holding potentials.

Acetylcholine↗
PubMed1980

Intracellular injection of t he catalytic subunit of cyclic AMP-dependent protein kinase simulates facilitation of transmitter release underlying behavioral sensitization in Aplysia.

It has been difficult to establish whether cyclic AMP-mediated protein phosphorylation in nerve cells plays a specific role in synaptic transmission. This difficulty can be overcome in higher invertebrates because their large neurons allow the injection of protein molecules into the cell. We have used intracellular injection to study whether protein phosphorylation is involved in the mechanism of sensitization, a simple form of learning. Sensitization of the gill-withdrawal reflex in Aplysia involves enhancement of transmitter release by presynaptic facilitation at a particular set of synaptic connections between identified sensory neurons and their follower cells. We have found that injection of the catalytic subunit of cyclic AMP-dependent protein kinase (ATP:protein phosphotransferase, EC 2.7.1.37) purified from bovine heart mimics the action of the natural transmitter and of serotonin, the putative transmitter, by simulating the physiological changes that accompany presynaptic facilitation. Intracellular injection of the kinase into a sensory cell (i) broadens the action potential in the presence of tetraethylammonium, indicating an increase in Ca2+ current, (ii) decreases the input conductance of the cell, presumably as a result of a decrease in the K+ current, and (iii) increases the amount of transmitter released by terminals of the sensory cell onto follower neurons.

Animals↗
PubMed1980

Presynaptic inhibition in Aplysia involves a decrease in the Ca2+ current of the presynaptic neuron.

By voltage clamping presynaptic cell L10 and using pharmacologic separation techniques, we have analyzed the specific ionic currents in the presynaptic cell that correlate with presynaptic inhibition while assaying transmitter release with intracellular recordings from postsynaptic cells. We have found that presynaptic inhibition can be elicited in conditions in which the Na+ and the various K+ channels are pharmacologically blocked and depolarizing current pulses produce only an inward Ca2+ current. Both inward currents and tail currents at and above the K+ reversal potential were always less inward during presynaptic inhibition. The changes in conductance associated with presynaptic inhibition were voltage sensitive and paralleled the voltage sensitivity of the Ca2+ channel. We therefore conclude that presynaptic inhibition is caused by a direct transmitter-mediated decreased of presynaptic Ca2+-channel conductance.

Animals↗
PubMed1979

Sensitization in Aplysia: restoration of transmission in synapses inactivated by long-term habituation.

Long-term habituation of a simple withdrawal reflex in Aplysia leads to an inactivation of synaptic transmission between identified sensory and gill motor neurons that persists for more than 3 weeks. A single sensitizing stimulus rapidly reactivates both the depressed behavioral response and the inactivated synaptic transmission. Thus sensitization, a simple competitive form of learning, provides a mechanism whereby changing environmental demands can rapidly override the long-term memory of habituation.

Animals↗
PubMed1979

Ultrastructure of the synapses of sensory neurons that mediate the gill-withdrawal reflex in Aplysia.

We have identified the processes of mechanoreceptor sensory neurons by intracellular injection of horseradish peroxidase in order to study the structure of synapses which exhibit profound, behaviourally-relevant plasticity. These synapses are located at small, varicose expansions along or at the end of the fine, microtubule-containing neurites, and they are crowded with vesicles some of which are associated with the varicosity membrane at regions of membrane specialization morphologically equivalent to active zones described in other species. These active zones occur between pre- and postsynaptic processes at two varieties of apposition: a conventional flat one, and a more elaborate indented one. At indented appositions, the presynaptic varicosity is invaginated by a thin (less than 0.25 micrometer diameter) spine of variable length. The active zones of indented synapses have approximately twice the vesicle frequency of flat synapses, suggesting that indented synapses are more effective. Sensory neuron terminals are relatively uniform in their structure, having similar concentrations of vesicles and numbers of active zones, and the majority of the processes postsynaptic to them are less than 0.5 micrometer in diameter. These regularities, and the presence of two strikingly-different types of synaptic apposition, flat and indented, should facilitate structural comparisons of neurons from naive and behaviourally-modified animals. The possible dynamic interconversion of indented and flat appositions at the synaptic terminals of sensory neurons and its behavioural relevance are discussed.

Animals↗
PubMed1979

Quantitative analysis of relative contribution of central and peripheral neurons to gill-withdrawal reflex in Aplysia californica.

1. There is general agreement that the gill-withdrawal reflex elicited by weak tactile stimuli (less than 2 g to the siphon skin) is mediated almost entirely by the central nervous system (CNS) (13, 15, 18, 23). However, there was disagreement concerning the effects of moderate intensity (2--4 g) stimuli. Kupfermann et al. (18) found that the CNS mediates approximately 94% of the reflex elicited by moderate-intensity stimuli, whereas Peretz et al. (23) found that in this stimulus range, the amplitude of the reflex was, on average, unaltered when the CNS was removed. 2. To resolve this difference we first carried out pilot experiments in collaboration with B. Peretz, J. W. Jacklet, and K. Lukowiak using isolated mantle preparations, and then performed a systematic study using both the isolated mantle and intact animals. The main difference that seemed to account for the discrepancy in the results was the magnitude of the reflex response that was selected for study. Previous studies from this laboratory used a minimum-response criterion whereby only brisk and clearly observable responses of at least 35% maximum were examined. By contrast, Peretz et al. (23) examined all responses, even those that were extremely small. In addition, the two groups used different methods of stimulation so that stimulus intensities could not really be compared. 3. By comparing the effects of moderate-intensity stimuli in experiments with and without a response criterion in isolated mantle preparations, we found that when a minimal response-amplitude criterion is imposed the CNS mediates 90--95% of the gill-withdrawal reflex, whether it is elicited by the "tapper" stimulus used by Peretz et al. (23) or by the servo-controlled probe previously used in this laboratory. On the other hand, when no minimal response criterion is used and small responses are also examined, the response to the probe is still significantly reduced by 85% when the CNS is removed, whereas the reflex response to the tapper is more variable, sometimes increasing and other times decreasing with deganglionation. 4. We have also tested, in intact animals, the role of the CNS in mediating gill withdrawal. Water-jet stimuli were delivered to the siphon to elicit the reflex, as in previous behavioral studies. As was the case with probe stimulation in the isolated mantle preparation, in intact animals the CNS mediates 90% of the reflex evoked by moderate-intensity stimuli when a minimal response-amplitude criterion is imposed. 5. Our experiments indicate that, using response criteria and methods of stimulation, one can reliably study the reflex as predominantly mediated by the CNS.

Animals↗
PubMed1979

Cellular studies of peripheral neurons in siphon skin of Aplysia californica.

1. To account for the similarity in the kinetics of habituation between the central and peripheral components of siphon withdrawal, we have tested the idea (52) that each centrally located mechanoreceptor sensory neuron sends two branches to siphon motor neurons; one to centrally located siphon motor neurons and a collateral branch that remains in the periphery and innervates the peripheral siphon motor neurons. 2. We have found a group of peripheral siphon motor neurons and tested the connection onto these cells by central mechanoreceptors. In addition, we have defined by various electrophysiological and morphological criteria two general classes of peripheral neurons that lie along the course of the siphon nerve. 3. One class (type I) consists of only a single cell in each animal. This peripheral neuron typically has the largest cell body found lying along the siphon nerve and is the only peripheral nerve cell that appears white when viewed under epi-illumination. The type I neuron often has a highly regular firing pattern, which occurs in the absence of spontaneous synaptic input. The three-dimensional morphology of this neuron suggests a paucity of fine processes, most of which do not arborize and may terminate in the connective tissue sheath. Fine structural observations of the peripheral white cell have revealed the presence of large densecore granules. The peripheral type I neuron is similar in most of its electrophysiological and morphological properties to central neurons postulated to be neurosecretory. The peripheral white cell is, at present, the only peripheral neuron we can identify with certainty as a unique individual. 4. The second class (type II) of peripheral neurons are siphon motor neurons for the peripheral component of the siphon-withdrawal reflex. In contrast to the type I neurons, members of the second class of peripheral neurons possess smaller, more spherical cell bodies that have varying amounts of orange pigmentation and which give rise to a relatively well-developed and arborized dendritic tree. Type II neurons feature an irregular spontaneous firing pattern that is occasionally modulated by a rich spontaneous synaptic input. Peripheral siphon motor neurons have restricted motor fields that produce contraction of the mantle floor and the base of the siphon. Most of the type II neurons were found to be electrically coupled to one another. 5. The peripheral siphon motor neurons resemble the central siphon motor neurons in that they receive a collateral synapse from centrally located mechanoreceptor sensory neurons. This peripheral sensory-to-motor synapse exhibits the same kinetics of decrement as its central counterpart, both of which parallel behavioral habituation. 6. The rich mechanoreceptor input onto the relatively isolated dendritic trees of the peripheral siphon motor neurons provide a uniquely restricted neuropil to study the sensory-to-motor synapse. The peripheral motor neurons may, therefore, be a useful simple preparation for the cellular study of behavioral plasticity.

Animals↗
PubMed1978

Cellular analysis of long-term habituation of the gill-withdrawal reflex of Aplysia californica.

Long-term habituation training in Aplysia californica produces a profound depression in the efficacy of synaptic transmission between mechanoreceptor neurons and gill motor neurons. This depression persists for more than 3 weeks. Thus a critical synaptic site for plasticity underlying long-term habituation is the same as that for short-term habituation. For this simple form of learning, short- and long-term memory share a common locus and aspects of a common mechanism: synaptic depression.

Animals↗
PubMed1978

Stimulus-response relations and stability of mechanoreceptor and motor neurons mediating defensive gill-withdrawal reflex in Aplysia.

1. A weak or moderate-intensity tactile stimulus delivered to the siphon skin of Aplysia californica elicits a defensive reflex withdrawal of the gill and siphon into the mantle cavity. The reflex undergoes both short- and long-term habituation and sensitization and has, therefore, been used as a model system to examine various forms of learning. In this paper we describe studies of the response properties of the sensory and motor neurons of the reflex during repeated stimulation at rates that produce habituation. 2. The sensory neurons are slowly adapting mechanoreceptor cells whose frequency of discharge is a monotonic function of controlled-force punctate stimuli delivered to the skin. The majority of the stimulus-response relations could best be described by exponential functions. 3. We examined the stability of the sensory neuron responses in two ways; with punctate stimuli of varying intensity and with water jets of varying intensity. 4. With repeated punctate stimulation at rates which produce habituation in the intact animal the mechanoreceptor discharge showed no decrement. This stability was observed over a 10-fold range of intensities. 5. Weak or moderate intensity water-jet stimuli to the skin also gave stable responses but stronger stimuli caused the mechanoreceptor response to fatigue. 6. We examined the stability of the motor responses by using intracellular depolarizing current pulses to produce repetitive bursts of action potentials in gill motor neurons while monitoring the gill contractions with a strain gauge, photocell, or videotape recorder. The photocell and strain gauge were alternatively used in the same experiment. Gill contractions monitored with the photocell were stable, whereas those monitored by the strain gauge showed decrement. An independent measure of gill contraction, videotape recording, confirmed the results obtained with the photocell and showed that the gill contractions following repeated intracellular depolarization of the motor neurons were stable.

Action Potentials↗
PubMed1978

Contribution of individual mechanoreceptor sensory neurons to defensive gill-withdrawal reflex in Aplysia.

1. To evaluate the contribution which mechanoreceptor sensory neurons make to the defensive gill-withdrawal reflex we developed an isolated reflex preparation. We then reduced this isolated reflex to a microcircuit (consisting of a single sensory cell and single motor cell) so as to causally relate the contribution of individual cells to the expression and plastic properties of the behavior. 2. Mechanoreceptor neurons make significant contributions to the amplitude and duration of the complex PSP in the motor neurons. A single spike in a sensory neuron produces an EPSP in the motor neuron which accounts for 7-36% of the complex EPSP produced by weak tactile stimulation of the skin. 3. More than 50% of the synaptic input to the gill motor neurons appears to be monosynaptic. Perfusing the ganglion with solutions of high divalent cations reduced the motor neurons' complex PSP by only 40%. 4. The population response of the mechanoreceptors to a point stimulus can be simulated by repetitively firing a single sensory neuron. Firing a single sensory cell discharges the motor neuron and produces a gill contraction similar to that produced by a natural stimulus. 5. Mechanoreceptors make monosynaptic connections onto gill motor neurons which decrement with repeated stimulation paralleling the decrement of the complex PSP to punctate tactile stimulation of the skin. 6. The results indicate that the known neural elements may quantitatively account for most of the expression of the behavior and its short-term habituation.

Animals↗
PubMed1974

A quantal analysis of the synaptic depression underlying habituation of the gill-withdrawal reflex in Aplysia.

Habituation, one of the simplest behavioral paradigms for studying memory, has recently been examined on the cellular level in the gill-withdrawal reflex in the mollusc Aplysia and in the escape response in cray-fish. In both cases short-term habituation involved a decrease in excitatory synaptic transmission at the synapses between the sensory neurons and their central target cells. To analyze the mechanisms of the synaptic depression in Aplysia, we applied a quantal analysis to synaptic transmission between the sensory and motor neurons of the gill-withdrawal reflex. Our results indicate that short-term habituation results from a presynaptic mechanism: a decrease in the number of transmitter quanta released per impulse. The sensitivity of the postsynaptic receptor remains unaltered.

Action Potentials↗