strychnine and biocytin

Unitary excitatory (EPSP) and inhibitory (IPSP) postsynaptic potentials (PSPs) were evoked between neurons in Rexed's laminae (L)II-V of spinal slices from young hamsters (7-24 days old) at 27°C using paired whole cell recordings. Laminar differences in synaptic efficacy were observed: excitatory connections were more secure than inhibitory connections in LII and inhibitory linkages in LII were less reliable than those in LIII-V. A majority of connections displayed paired-pulse facilitation or depression. Depression was observed for both EPSPs and IPSPs, but facilitation was seen almost exclusively for IPSPs. There were no frequency-dependent shifts between facilitation and depression. Synaptic depression was associated with an increased failure rate and decreased PSP half-width for a majority of connections. However, there were no consistent changes in failure rate or PSP time course at facilitating connections. IPSPs evoked at high-failure synapses had consistently smaller amplitude and showed greater facilitation than low-failure connections. Facilitation at inhibitory connections was positively correlated with synaptic jitter and associated with a decrease in latency. At many connections, the paired-pulse ratio varied from trial to trial and depended on the amplitude of the first PSP; dependence was greater for inhibitory synapses than excitatory synapses. Paired-pulse ratios for connections onto neurons with rapidly adapting, "phasic" discharge to depolarizing current injection were significantly greater than for connections onto neurons with tonic discharge properties. These results are evidence of diversity in synaptic transmission between dorsal horn neurons, the nature of which may depend on the types of linkage, laminar location, and intrinsic firing properties of postsynaptic cells.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Animals, Newborn; Bicuculline; Cricetinae; Electric Stimulation; Excitatory Amino Acid Antagonists; Female; GABA-A Receptor Antagonists; Glycine Agents; In Vitro Techniques; Lysine; Male; Mesocricetus; Neural Pathways; Patch-Clamp Techniques; Posterior Horn Cells; Reaction Time; Spinal Cord; Strychnine; Synapses; Synaptic Transmission; Time Factors

Proprioceptive sensory signals inform the CNS of the consequences of motor acts, but effective motor planning involves internal neural systems capable of anticipating actual sensory feedback. Just where and how predictive systems exert their influence remains poorly understood. We explored the possibility that spinocerebellar neurons that convey proprioceptive sensory information also integrate information from cortical command systems. Analysis of the circuitry and physiology of identified dorsal spinocerebellar tract neurons in mouse spinal cord revealed distinct populations of Clarke's column neurons that received direct excitatory and/or indirect inhibitory inputs from descending corticospinal axons. The convergence of these descending inhibitory and excitatory inputs to Clarke's column neurons established local spinal circuits with the capacity to mark or modulate incoming proprioceptive input. Together, our genetic, anatomical and physiological results indicate that Clarke's column spinocerebellar neurons nucleate local spinal corollary circuits that are relevant to motor planning and evaluation.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Animals, Newborn; beta-Galactosidase; Bicuculline; Cerebellum; Cholera Toxin; Electric Stimulation; Estrogen Antagonists; Excitatory Amino Acid Antagonists; Feedback, Sensory; GABA Antagonists; Gene Expression Regulation; Glial Cell Line-Derived Neurotrophic Factor; Glycine Agents; Glycine Plasma Membrane Transport Proteins; Green Fluorescent Proteins; Homeodomain Proteins; Lysine; Membrane Potentials; Mice; Mice, Transgenic; Models, Neurological; Nerve Net; Neural Inhibition; Neural Pathways; Parvalbumins; Patch-Clamp Techniques; Protein Kinase C; RNA, Messenger; Sensory Receptor Cells; Spinal Cord; Stilbamidines; Strychnine; Tamoxifen; tau Proteins; Transcription Factors; Vesicular Glutamate Transport Protein 1

Responses of on-center starburst amacrine cells to steady light stimuli were recorded in the dark-adapted mouse retina. The response to spots of dim white light appear to show two components, an initial peak that correspond to the onset of the light stimulus and a series of oscillations that ride on top of the initial peak relaxation. The frequency of oscillations during light stimulation was three time higher than the frequency of spontaneous oscillations recorded in the dark. The light-evoked responses in starburst cells were exclusively dependent on the release of glutamate likely from presynaptic bipolar axon terminals and the binding of glutamate to AMPA/kainate receptors because they were blocked by 6-cyano-7-nitroquinoxalene-2,3-dione. The synaptic pathway responsible for the light responses was blocked by AP4, an agonist of metabotropic glutamate receptors that hyperpolarize on-center bipolar cells on activation. Light responses were inhibited by the calcium channel blockers cadmium ions and nifedipine, suggesting that the release of glutamate was calcium dependent. The oscillatory component of the response was specifically inhibited by blocking the glutamate transporter with d-threo-beta-benzyloxyaspartic acid, suggesting that glutamate reuptake is necessary for the oscillatory release. GABAergic antagonists bicuculline, SR 95531, and picrotoxin increased the amplitude of the initial peak while they inhibit the frequency of oscillations. TTX had a similar effect. Strychnine, the blocker of glycine receptors did not affect the initial peak but strongly decreased the oscillations frequency. These inhibitory inputs onto the bipolar axon terminals shape and synchronize the oscillatory component.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Amacrine Cells; Animals; Aspartic Acid; Biological Clocks; Cadmium Chloride; Calcium Channel Blockers; Electric Stimulation; Excitatory Amino Acid Antagonists; Glycine Agents; In Vitro Techniques; Light; Lysine; Membrane Potentials; Mice; Mice, Inbred C57BL; Nifedipine; Patch-Clamp Techniques; Photic Stimulation; Retina; Strychnine; Synapses

The principal cells of the chick tangential nucleus are vestibular nucleus neurons participating in the vestibuloocular and vestibulocollic reflexes. In birds and mammals, spontaneous and stimulus-evoked firing of action potentials is essential for vestibular nucleus neurons to generate mature vestibular reflex activity. The emergence of spike-firing pattern and the underlying ion channels were studied in morphologically-identified principal cells using whole-cell patch-clamp recordings from brain slices of late-term embryos (embryonic day 16) and hatchling chickens (hatching day 1 and hatching day 5). Spontaneous spike activity emerged around the perinatal period, since at embryonic day 16 none of the principal cells generated spontaneous action potentials. However, at hatching day 1, 50% of the cells fired spontaneously (range, 3 to 32 spikes/s), which depended on synaptic transmission in most cells. By hatching day 5, 80% of the principal cells could fire action potentials spontaneously (range, 5 to 80 spikes/s), and this activity was independent of synaptic transmission and showed faster kinetics than at hatching day 1. Repetitive firing in response to depolarizing pulses appeared in the principal cells starting around embryonic day 16, when <20% of the neurons fired repetitively. However, almost 90% of the principal cells exhibited repetitive firing on depolarization at hatching day 1, and 100% by hatching day 5. From embryonic day 16 to hatching day 5, the gain for evoked spike firing increased almost 10-fold. At hatching day 5, a persistent sodium channel was essential for the generation of spontaneous spike activity, while a small conductance, calcium-dependent potassium current modulated both the spontaneous and evoked spike firing activity. Altogether, these in vitro studies showed that during the perinatal period, the principal cells switched from displaying no spontaneous spike activity at resting membrane potential and generating one spike on depolarization to the tonic firing of spontaneous and evoked action potentials.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Action Potentials; Animals; Apamin; Bicuculline; Cesium; Chick Embryo; Chlorides; Dose-Response Relationship, Radiation; Drug Combinations; Electric Stimulation; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; GABA Antagonists; In Vitro Techniques; Lysine; Neurons; Sodium Channel Blockers; Strychnine; Tetrodotoxin; Vestibular Nuclei

The contributions of the hyperpolarization-activated current, I(h), to generation of rhythmic activities are well described for various central neurons, particularly in thalamocortical circuits. In the present study, we investigated effects of a general anesthetic, propofol, on native I(h) in neurons of thalamus and cortex and on the corresponding cloned HCN channel subunits. Whole cell voltage-clamp recordings from mouse brain slices identified neuronal I(h) currents with fast activation kinetics in neocortical pyramidal neurons and with slower kinetics in thalamocortical relay cells. Propofol inhibited the fast-activating I(h) in cortical neurons at a clinically relevant concentration (5 microM); inhibition of I(h) involved a hyperpolarizing shift in half-activation voltage (DeltaV1/2 approximately -9 mV) and a decrease in maximal available current (approximately 36% inhibition, measured at -120 mV). With the slower form of I(h) expressed in thalamocortical neurons, propofol had no effect on current activation or amplitude. In heterologous expression systems, 5 muM propofol caused a large shift in V1/2 and decrease in current amplitude in homomeric HCN1 and linked heteromeric HCN1-HCN2 channels, both of which activate with fast kinetics but did not affect V1/2 or current amplitude of slowly activating homomeric HCN2 channels. With GABA(A) and glycine receptor channels blocked, propofol caused membrane hyperpolarization and suppressed action potential discharge in cortical neurons; these effects were occluded by the I(h) blocker, ZD-7288. In summary, these data indicate that propofol selectively inhibits HCN channels containing HCN1 subunits, such as those that mediate I(h) in cortical pyramidal neurons-and they suggest that anesthetic actions of propofol may involve inhibition of cortical neurons and perhaps other HCN1-expressing cells.

Topics: Animals; Animals, Newborn; Anticonvulsants; Bicuculline; Cerebral Cortex; Cyclic Nucleotide-Gated Cation Channels; Dose-Response Relationship, Drug; Dose-Response Relationship, Radiation; Drug Interactions; Electric Stimulation; Female; GABA Antagonists; Glycine Agents; Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; In Vitro Techniques; Ion Channel Gating; Ion Channels; Lysine; Male; Membrane Potentials; Mice; Neural Inhibition; Neural Pathways; Oocytes; Patch-Clamp Techniques; Potassium Channels; Propofol; Pyramidal Cells; Pyrimidines; Rats; Strychnine; Thalamus; Time Factors; Xenopus

In vitro models have frequently been employed to investigate the specificity of the formation of axonal projections during both development and regeneration. Such studies demonstrated pathway, target, and laminar specificity, yet they did not tackle the problem of topography. Here, we addressed the issue of regeneration of spatial specificity at the topographic level by lesioning a precisely organized projection from the auditory system of neonatal rats in organotypic slice culture and by analyzing regeneration capacity. Lesioning had no effect on the survival of axotomized neurons or the structure of the auditory nuclei. Anterograde and retrograde biocytin tracing demonstrated that the projection regenerated topographically at the supracellular level. Whole-cell patch-clamp recordings revealed that the regenerated projection was functional. Topographic regeneration was not impaired by blocking spike activity with tetrodotoxin or glycinergic transmission with strychnine. However, if lesioning was performed after the slices had been incubated for 1 week, regeneration capacity was lost despite good survival of neurons. The loss of the regeneration capacity in vitro occurs at a developmental stage that corresponds to the age when the capacity for axonal reorganization is lost in vivo. We conclude that the developmental processes occurring in vivo and in vitro are comparable in this system, which is why we think that essential aspects of the loss of regeneration capacity may be addressed with our culture model in the future.

Topics: Animals; Auditory Pathways; Axons; Brain Stem; Evoked Potentials; gamma-Aminobutyric Acid; Glycine; Lysine; Nerve Regeneration; Neurons; Olivary Nucleus; Organ Culture Techniques; Patch-Clamp Techniques; Rats; Rats, Sprague-Dawley; Reaction Time; Strychnine; Synapses; Tetrodotoxin