Epileptiform activity in microcultures containing small numbers of hippocampal neurons

Segal, Michael M.; Furshpan, E. J.

doi:10.1152/jn.1990.64.5.1390

Cited by 137 publications

(110 citation statements)

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“…It is known that synaptic interactions between cultured hippocampal neurons can be very strong, especially in low density cultures, presumably due to multiple synaptic contacts between pairs of neurons (Bekkers and Stevens, 1995;Segal and Furshpan, 1990;Wilcox et al, 1994). The spiking of a single neuron can be sufficient to drive its postsynaptic targets above the threshold for spiking (Segal and Furshpan, 1990).…”

Section: Discussionmentioning

confidence: 99%

“…The spiking of a single neuron can be sufficient to drive its postsynaptic targets above the threshold for spiking (Segal and Furshpan, 1990). Therefore, complex synaptic responses due to the superposition of many polysynaptic pathways are often observed.…”

Section: Discussionmentioning

confidence: 99%

“…Then neurons are plated at low density. The microdots physically limit the neurons' growth so that some microislands end up with just a few neurons or even just two Stevens, 1991, 1995;Segal and Furshpan, 1990). Within such a microisland, the probability of connection is high, so that it is straightforward to find connected pairs of neurons (Kaplan et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

“…Furthermore, pairs of neurons on microislands tend to be strongly coupled, probably by multiple synaptic contacts (Segal and Furshpan, 1990). Evoked postsynaptic currents are typically hundreds of picoamperes or more, while spontaneous postsynaptic currents ("mini"s) are tens of picoamperes or less (Wilcox et al, 1994).…”

Section: Introductionmentioning

confidence: 99%

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Laser-evoked synaptic transmission in cultured hippocampal neurons expressing channelrhodopsin-2 delivered by adeno-associated virus

Wang¹,

Hasan²,

Seung³

2009

Journal of Neuroscience Methods

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We present a method for studying synaptic transmission in mass cultures of dissociated hippocampal neurons based on patch clamp recording combined with laser stimulation of neurons expressing Channelrhodopsin-2 (ChR2). Our goal was to use the high spatial resolution of laser illumination to come as close as possible to the ideal of identifying monosynaptically coupled pairs of neurons, which is conventionally done using microisland rather than mass cultures. Using recombinant adenoassociated virus (rAAV) to deliver the ChR2 gene, we focused on the time period between 14 and 20 days in vitro, during which expression levels are high, and spontaneous bursting activity has not yet started. Stimulation by wide-field illumination is sufficient to make the majority of ChR2-expressing neurons spike. Stimulation with a laser spot at least 10 μm in diameter also produces action potentials, but in a reduced fraction of neurons. We studied synaptic transmission by voltageclamping a neuron with low expression of ChR2 and scanning a 40 μm laser spot at surrounding locations. Responses were observed to stimulation at a subset of locations in the culture, indicating spatial localization of stimulation. Pharmacological means were used to identify responses that were synaptic. Many responses were of smaller amplitude than those typically found in microisland cultures. We were unable to find an entirely reliable criterion for distinguishing between monosynaptic and polysynaptic responses. However, we propose that postsynaptic currents with Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. HHMI Author ManuscriptHHMI Author Manuscript HHMI Author Manuscript small amplitudes, simple shapes, and latencies not much greater than 8 msec are reasonable candidates for monosynaptic interactions.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Laser-evoked synaptic transmission in cultured hippocampal neurons expressing channelrhodopsin-2 delivered by adeno-associated virus

Wang¹,

Hasan²,

Seung³

2009

Journal of Neuroscience Methods

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“…The osmolality was 290-300 mOsmol/kg. TTX and kynurenic acid were included to help suppress Na + spikes and glutamate receptor activation, as in other studies (e.g., Segal and Furshpan, 1990). After this incubation, the retina and adhering filter were placed onto the support screen of a disc filter holder.…”

Section: Removing the Outer Retinamentioning

confidence: 99%

Dissociation of retinal ganglion cells without enzymes

Hayashida

Partida

Ishida

2004

Journal of Neuroscience Methods

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We describe here methods for dissociating retinal ganglion cells from adult goldfish and rat without proteolytic enzymes, and show responses of ganglion cells isolated this way to step-wise voltage changes and fluctuating current injections. Taking advantage of the laminar organization of vertebrate retinas, photoreceptors and other cells were lifted away from the distal side of freshly isolated goldfish retinas, after contact with pieces of membrane filter. Likewise, cells were sliced away from the distal side of freshly isolated rat retinas, after these adhered to a membrane filter. The remaining portions of retina were incubated in an enzyme-free, low Ca 2+ solution, and triturated. After aliquots of the resulting cell suspension were plated, ganglion cells could be identified by dye retrogradely transported via the optic nerve. These cells showed no obvious morphological degeneration for several days of culture. Perforated-patch whole-cell recordings showed that the goldfish ganglion cells spike tonically in response to depolarizing constant current injections, that these spikes are temporally precise in response to fluctuating current injections, and that the largest voltage-gated Na + currents of these cells were larger than those of ganglion cells isolated with a neutral protease.

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Enantioselective modulation of GABAergic synaptic transmission by steroids and benz[e]indenes in hippocampal microcultures

Zorumski

Mennerick

Covey

1998

Synapse

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The effects of enantiomers of the neurosteroid analogues, 3alpha-hydroxy-5alpha-pregnan-20-one (DHP) and 3alpha-hydroxy-5alpha-androstane-17beta-carbonitrile (ACN), and the benz[e]indene, BI-1, on synaptic currents were examined in microcultures of rat hippocampal neurons. Over the range of 0.1-10 microM, the (+)-enantiomers were more potent and effective than their (-)-enantiomeric counterparts in enhancing gamma-aminobutyric acid (GABA)A receptor-mediated evoked synaptic currents. The (+)-enantiomers had small effects on peak currents, but slowed the decay of inhibitory synaptic currents, resulting in 2-3-fold increases in charge transfer during inhibitory synaptic events at 10 microM. Similar prolongations of spontaneous miniature inhibitory postsynaptic currents (IPSCs) and responses to brief GABA pulses to outside-out patches suggest that the prolongations of evoked synaptic currents result primarily from postsynaptic effects. In contrast, the (-)-enantiomers had little effect on evoked IPSCs at concentrations < or = 1 microM, but enhanced inhibitory transmission at 10 microM. At concentrations < or = 1 microM, neither the (+)- nor (-)-enantiomers altered glutamate-mediated excitatory synaptic currents. At 10 microM, (+)-DHP and (+)-ACN depressed excitatory responses in a bicuculline-sensitive fashion, suggesting that direct chloride channel gating by the steroids contributed to the depression. These data indicate that certain steroids and benz[e]indenes augment inhibitory synaptic transmission enantioselectively and provide strong support for the hypothesis that steroids act at specific sites on synaptic GABA(A) receptors rather than via alteration of membrane lipids.

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Epileptiform activity in microcultures containing small numbers of hippocampal neurons

Cited by 137 publications

References 0 publications

Laser-evoked synaptic transmission in cultured hippocampal neurons expressing channelrhodopsin-2 delivered by adeno-associated virus

Laser-evoked synaptic transmission in cultured hippocampal neurons expressing channelrhodopsin-2 delivered by adeno-associated virus

Dissociation of retinal ganglion cells without enzymes

Enantioselective modulation of GABAergic synaptic transmission by steroids and benz[e]indenes in hippocampal microcultures

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