Identified Aplysia neurons form specific chemical synapses in culture

Camardo, Joseph S.; Proshansky, Eric; Schacher, Samuel

doi:10.1523/jneurosci.03-12-02614.1983

Cited by 76 publications

(58 citation statements)

References 29 publications

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“…4D). However, with the cell poised at hyperpolarizing potentials, either by the injection of current or the addition of acetylcholine (16), the rate of depolarization in the processes was substantially less than that in the soma (Fig. 4 F and G).…”

Section: Resultsmentioning

confidence: 99%

Noninvasive detection of changes in membrane potential in cultured neurons by light scattering.

Stepnoski

LaPorta

Raccuia-Behling

et al. 1991

Proc. Natl. Acad. Sci. U.S.A.

225

159

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We report a procedure to detect electrical activity in cultured neurons by changes in their intrinsic opticalproperties. Using dark-field microscopy to detect scattered light, we observe an optical signal that is linearly proportional to the change in the membrane potential. Action potentials can be recorded without signal averaging. We use the dark-field method to show that there are substantial time delays between activity in the soma and in fine distal processes of identified Aplyswa neurons. The biophysical basis for the change in optical properties of the neuron was deduced from measurements of the angular distribution of scattered laser light. An analysis of the data indicates that the radial component of the index of refraction of the membrane increases and the tangential components decrease concomitant with an increase in membrane potential. This is suggestive of a rapid reorientation of dipoles in the membrane during an action potential. Here we reexamine the coupling between the optical properties of neurons and changes in their transmembrane potential. We are motivated by the desire to elucidate a biophysical mechanism that mediates this coupling and by the need to establish a noninvasive means to probe the electrical activity of neurons. Our model system consisted of identified invertebrate neurons. These cells play an increasing role in the study of synaptic plasticity and in the study of emergent properties of small networks. In the present context, they allow us to probe optical changes in a variety ofcell types, old versus new outgrowth, and thick versus fine processes.EXPERIMENTAL METHODS Cell Culture and Electrophysiology. Juvenile Aplysia californica (2-10 g; stage 12), raised in mariculture (18) purchased from the University of Miami (Florida). Identified neurons in the abdominal and cerebral ganglia were located, microdissected from the ganglion, and maintained in cell culture, as described (6, 7). The cells consist of a soma and one or two axonal stumps at the time of isolation (Fig. 1A). They generate an extensive array of processes after 2-3 days in vitro (Fig. 1B). We estimated the area of cell membrane in the processes as described (8). Standard current-clamp and voltage-clamp methods were used for intracellular stimulation and recording.Dark-Field Microscopy. The optical system was centered around an upright microscope (UEM; Zeiss) with a x40 (0.75 n.a.) water-immersion lens (no. 46-17-02; Zeiss) and a (0.80-0.95 n.a.) dark-field illuminator (no. 46-55-05; Zeiss) (see Fig. 2A Inset). The cells were plated on a glass coverslip that formed the bottom of a 50-mm Petri dish with a hole drilled in the bottom. For all records shown, except those in Fig. 4B, the light source was a 100-W quartz tungsten lamp (Xenophot HLX; Osram, Berlin) that was filtered by 1 cm of heatabsorbing glass. The maximum power at the preparation was 1 x 10-3 W and the relative fluctuations (peak to peak) in the intensity were typically -3 x 10-5 in the frequency range 1-300 Hz. For measurements of the wavel...

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Section: Resultsmentioning

confidence: 99%

Noninvasive detection of changes in membrane potential in cultured neurons by light scattering.

Stepnoski

LaPorta

Raccuia-Behling

et al. 1991

Proc. Natl. Acad. Sci. U.S.A.

225

159

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“…These plastic properties, especially homosynaptic depression, are specific to the LE-L7 circuit. Other Aplysia cells that form chemical connections in vitro show stable PSP amplitudes, with stimulation rates that depress the LE-L7 connection (Camardo et al, 1983;Schacher et al, 1985). Heterosynaptic facilitation of LE-L7 connections can also be produced by other cells.…”

Section: Discussionmentioning

confidence: 99%

Synaptic plasticity in vitro: cell culture of identified Aplysia neurons mediating short-term habituation and sensitization

Rayport¹,

Schacher²

1986

J. Neurosci.

Self Cite

174

124

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The gill withdrawal reflex of the marine mollusk, Ap&u californica, shows habituation and sensitization, two simple forms of learning. In order to extend the cellular studies on synaptic plasticity underlying the changes in the reflex behavior, and to explore further the development of synaptic plasticity during synapse formation, we have sought to establish the neural circuit of the gill withdrawal reflex in vitro. We report here the reconstruction of the elementary gill withdrawal circuit in cell culture and find that the cells show short-term homosynaptic depression and heterosynaptic facilitation, the cellular mechanisms of habituation and sensitization, respectively.The gill withdrawal reflex of the marine snail, Aplysia calijbrnica, is a simple defensive reflex mediated by a population of about 24 sensory cells, six motor cells, and several interneurons (Kandel, 1979). The reflex undergoes two forms of nonassociative learning, habituation and sensitization, as well as classical conditioning Kandel and Schwartz, 1982). The elementary cellular substrate underlying these forms of learning consists of sensory neurons that monosynaptically connect with a motor cell and a facilitatory neuron that synapses onto the presynaptic terminals of the sensory neurons (Bailey et al., 1979(Bailey et al., , 1981. Analysis of these forms of learning at the cellular level indicates that stimulus experience modulates transmitter release from the terminals of the sensory neurons Klein et al., 1980). Habituation of the reflex is mediated by homosynaptic depression, a decrease in transmitter release from the sensory neuron presynaptic terminals with repeated stimulation . Sensitization is based on heterosynaptic (presynaptic) facilitation, an increase in transmitter release from sensory neurons due in part to a reduction in a K+ current, and a consequent increased CaZ+ influx Klein and Kandel, 1978;Klein et al., 1982). Sensitization in the intact nervous system is mediated either by activity in a number of facilitator neurons, some of which are identified, or by the exogenous facilitatory neurotransmitters, namely, serotonin (5-HT) or the small molReceived Mar. 22, 1985; revised May 28, 1985; accepted June 18, 1985. This work was supported by National Science Foundation Grant 15948 to S.S. and National Institutes of Health Grant GM 32099, which supports the mariculture facility at the Marine Biological Laboratory. We wish to thank R. Woolley and L. Katz for their technical assistance, T. Capo and S. Per&t for raising the animals used in this study, and H. Ayers for typing the manuscript. We also thank Drs. E. Kandel, V. Castellucci, and M. Klein for their comments on earlier drafts of the manuscript.Correspondence should be addressed to Dr. (Abrams et al., 1984;Brunelli et al., 1976; Hawkins et al., 1981b).To gain direct access to the synapses mediating the learning exhibited by this reflex, and to enable us to explore the developmental processes involved in the acquisition of plastic properties by newly formed synapses, we ...

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“…The identification of cultured neurons is crucial because synaptic properties depend on the various combinations of presynaptic and postsynaptic neurons, and these combination-specific properties ultimately determine the functional role of the neural circuitry. In invertebrates, synapses were formed between identified neurons in culture (16,17). In mammals, synaptic formation between sensory neurons ofdorsal root ganglia and spinal neurons have been reported (18,19); however, spinal neurons were not further identified.…”

Section: Discussionmentioning

confidence: 99%

Cerebellar granule cells in culture: monosynaptic connections with Purkinje cells and ionic currents.

Hirano¹,

Kubo²,

Wu³

1986

Proc. Natl. Acad. Sci. U.S.A.

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Identified Aplysia neurons form specific chemical synapses in culture

Cited by 76 publications

References 29 publications

Noninvasive detection of changes in membrane potential in cultured neurons by light scattering.

Noninvasive detection of changes in membrane potential in cultured neurons by light scattering.

Synaptic plasticity in vitro: cell culture of identified Aplysia neurons mediating short-term habituation and sensitization

Cerebellar granule cells in culture: monosynaptic connections with Purkinje cells and ionic currents.

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