Hepatic encephalopathy (HE) results from impaired detoxification capacity of the liver leading to... more Hepatic encephalopathy (HE) results from impaired detoxification capacity of the liver leading to accumulation of various substances that may impair the function of the central nervous system (CNS).1—5
Spontaneously active neuronal networks grown from embryonic murine frontal cortex on substrate in... more Spontaneously active neuronal networks grown from embryonic murine frontal cortex on substrate integrated electrode arrays with 64 recording sites were used to assess acute neurobiological and toxic effects of a series of seven symmetrical, bifunctional alkylene-linked bis-thiocarbonate compounds designed to possess anticholinesterase activity. Acute functional neurotoxicity in the absence of cytotoxicity was defined as total collapse of spontaneous activity. All of the compounds were characterized as mixed inhibitors of AChE, with K(i)'s in the 10(-7)-10(-6) M range. The neuronal network assays revealed high repeatability for each compound, but surprisingly diverse effects among these closely related compounds. Six of the seven compounds produced changes in network activity at concentrations of 10-350 microM. Three of the compounds were excitatory, two were biphasic (excitatory at lower concentrations, inhibitory at higher), and one was solely inhibitory. Two of the inhibitory compounds produced irreversible inhibition of activity. Responses of cortical cultures to eserine were compared to the effects produced by the test compounds, with only one of seven providing a close match to the eserine profile. Matching of response patterns allows the classification of new drugs according to their response similarity to well-characterized agents. Spontaneously active neuronal networks reflect the interactions of multiple neurotransmitter and receptor systems, and can reveal unexpected side effects due to secondary binding. Utilizing such networks holds the promise of greater research efficiency through a more rapid recognition of physiological tissue responses.
THE NATURE AND IMPORTANCE of endogenous protective or recovery responses following sublethal acut... more THE NATURE AND IMPORTANCE of endogenous protective or recovery responses following sublethal acute physical injury have received little attention in investigations of CNS physical injury. The symposium focused attention on this important area of trauma research. An ...
An in vitro investigation was undertaken to provide information regarding the effectiveness of me... more An in vitro investigation was undertaken to provide information regarding the effectiveness of methylprednisolone sodium succinate (MPSS) as a treatment for the primary mechanical injury of spinal cord (SC) trauma. Exposure of uninjured mouse SC cells to MPSS for 24 h caused neuronal stress when the concentration exceeded 150 micrograms/mL; neuronal death occurred at concentrations above 600 micrograms/mL. The concentration range for MPSS protection of SC neurons subjected to a defined physical injury (laser microbeam transection of a primary dendrite 100 microns from the perikaryon) was very narrow: survival in the 30 micrograms/mL group differed significantly from the untreated control group (68.5% +/- 14.1 vs. 47.1% +/- 14.1), treatment with 20 or 60 micrograms/mL MPSS did not increase survival, and treatment with 100 micrograms/mL MPSS accelerated ultrastructural deterioration and increased the likelihood of death. Enhanced survival of lesioned neurons was observed when 30 micrograms/mL MPSS was applied within 15 min of dendrotomy but not when MPSS was administered 2 h after lesioning. Multimicroelectrode plate (MMEP) studies of SC network electrical activity indicated that MPSS associated readily with neuronal membranes. This finding was consistent with the hypothesis that MPSS may protect lesioned neurons by stabilizing damaged membranes, enhancing lesion resealing, and limiting the spread of ion-mediated damage. However, comparisons of neurite die-back 24 h after dendrotomy found no significant difference between MPSS-treated and control neurons. Application of 30 or 100 micrograms/mL MPSS increased the spontaneous burst activity of SC networks grown on MMEPs, however, there was no evidence that the increased excitability at these concentrations was the result of specific actions of MPSS on GABA or NMDA synapses.
Implanting electrical devices in the nervous system to treat neural diseases is becoming very com... more Implanting electrical devices in the nervous system to treat neural diseases is becoming very common. The success of these brain-machine interfaces depends on the electrodes that come into contact with the neural tissue. Here we show that conventional tungsten and stainless steel wire electrodes can be coated with carbon nanotubes using electrochemical techniques under ambient conditions. The carbon nanotube coating enhanced both recording and electrical stimulation of neurons in culture, rats and monkeys by decreasing the electrode impedance and increasing charge transfer. Carbon nanotube-coated electrodes are expected to improve current electrophysiological techniques and to facilitate the development of long-lasting brain-machine interface devices.
Prolonged (2-6 h) cooling of monolayer cultures of dissociated murine spinal cord at temperatures... more Prolonged (2-6 h) cooling of monolayer cultures of dissociated murine spinal cord at temperatures below 17 degrees C caused pronounced swelling of neuronal perikarya and dendrites. The numbers of swollen neurons in a culture increased as the temperature was reduced, and at 7 degrees C-10 degrees C all of the neurons were swollen. On rewarming the cultures to 37 degrees C, the majority of the swollen neurons died (up to 74% at 10 degrees C). Glial cells were not affected. Addition of the NMDA antagonists D-2-amino-5-phosphonovalerate (DAPV, 100 microM), ketamine (100 microM), and dibenzocyclohepteneimine (MK801, 10 microM) to spinal cord cultures before lowering the temperature to 10 degrees C minimized the dendrosomatic swelling and reduced neuronal mortality from 74% to 10%. These data show a surprising sensitivity of some neurons to nonfreezing low temperatures and suggest direct involvement of the NMDA receptor in hypothermia-related neuronal death.
Bodian and Nissl procedures were combined to stain dissociated mouse spinal cord cells cultured o... more Bodian and Nissl procedures were combined to stain dissociated mouse spinal cord cells cultured on coverslips. The Bodian technique stains fine neuronal processes in great detail as well as an intracellular fibrillar network concentrated around the nucleus and in proximal neurites. The Nissl stain clearly delimits neuronal cytoplasm in somata and in large dendrites. A combination of these techniques allows the simultaneous depiction of neuronal perikarya and all afferent and efferent processes. Costaining with little background staining by either procedure suggests high specificity for neurons. This procedure could be exploited for routine network analysis of cultured neurons.
The Journal of neuroscience : the official journal of the Society for Neuroscience, 1986
Laser-deinsulated, printed-circuit electrodes integrated into the floor of culture chambers have ... more Laser-deinsulated, printed-circuit electrodes integrated into the floor of culture chambers have been used to monitor the spontaneous activity of mouse spinal monolayer cell cultures. This technique has allowed a multisite analysis of activity over long periods of time in closed chambers. In 63 cultures investigated 3-5 weeks after seeding, 89% included single- or multiunit bursting. Based on a subset of 40 cultures in which all electrodes were sequentially scanned, bursting was found on 41% of the active electrodes (approximately 38% of all units monitored). A total of 35% of the electrodes monitoring spontaneous bursting activity revealed rhythmic sequences that were usually coupled among multiple electrodes. Although most of this coupling was in-phase, three out of 40 cultures exhibited antiphasic bursting. In all cases where coupling was observed, each electrode monitored different burst compositions, demonstrating that the activity was generated by different units. Some rhythmi...
Cultured spinal cord networks grown on microelectrode arrays display complex patterns of spontane... more Cultured spinal cord networks grown on microelectrode arrays display complex patterns of spontaneous burst and spike activity. During disinhibition with bicuculline and strychnine, synchronized burst patterns routinely emerge. However, the variability of both intra- and interculture burst periods and durations are typically large under these conditions. As a further step in simplification of synaptic interactions, we blocked excitatory AMPA synapses with 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzoquinoxaline-7-sulphonamide (NBQX), resulting in network activity mediated through the N-methyl-D-aspartate (NMDA) receptor (NMDA(ONLY)). This activity was APV sensitive. The oscillation under NMDA(ONLY) conditions at 37 degrees C was characterized by a period of 2.9 +/- 0.3 s (16 separate cultures). More than 98% of all neurons recorded participated in this highly rhythmic activity. The temporal coefficients of variation, reflecting the rhythmic nature of the oscillation, were 3.7, 4.7, and 4...
The Journal of neuroscience : the official journal of the Society for Neuroscience, 1983
The exposure of neuronal and glial cell processes to a large number (up to 300) of 12-nsec laser ... more The exposure of neuronal and glial cell processes to a large number (up to 300) of 12-nsec laser pulses at a wavelength of 337 nm and energy densities below the threshold for nonlinear absorption results in a gradual, gentle process transection in the laser focus. Within 10 to 20 sec after cessation of firing, the process pinches in the target area. During this time, mitochondria become swollen and bleached, the plasma membrane develops an obvious tautness, microtubules disappear, and organelles accumulate to either side of the process constriction. Depending on the irradiation parameters, a local pinching may proceed to a transection in about 30 sec or it may reverse to yield a normal-appearing process in approximately 5 min. Severe process pinching is accompanied by a sudden depolarization that may last for 2 to 5 min and is usually followed by a repolarization to the original resting potential even if the process has transected. Spiral retraction of cut processes and cytoplasmic ...
Hepatic encephalopathy (HE) results from impaired detoxification capacity of the liver leading to... more Hepatic encephalopathy (HE) results from impaired detoxification capacity of the liver leading to accumulation of various substances that may impair the function of the central nervous system (CNS).1—5
Spontaneously active neuronal networks grown from embryonic murine frontal cortex on substrate in... more Spontaneously active neuronal networks grown from embryonic murine frontal cortex on substrate integrated electrode arrays with 64 recording sites were used to assess acute neurobiological and toxic effects of a series of seven symmetrical, bifunctional alkylene-linked bis-thiocarbonate compounds designed to possess anticholinesterase activity. Acute functional neurotoxicity in the absence of cytotoxicity was defined as total collapse of spontaneous activity. All of the compounds were characterized as mixed inhibitors of AChE, with K(i)'s in the 10(-7)-10(-6) M range. The neuronal network assays revealed high repeatability for each compound, but surprisingly diverse effects among these closely related compounds. Six of the seven compounds produced changes in network activity at concentrations of 10-350 microM. Three of the compounds were excitatory, two were biphasic (excitatory at lower concentrations, inhibitory at higher), and one was solely inhibitory. Two of the inhibitory compounds produced irreversible inhibition of activity. Responses of cortical cultures to eserine were compared to the effects produced by the test compounds, with only one of seven providing a close match to the eserine profile. Matching of response patterns allows the classification of new drugs according to their response similarity to well-characterized agents. Spontaneously active neuronal networks reflect the interactions of multiple neurotransmitter and receptor systems, and can reveal unexpected side effects due to secondary binding. Utilizing such networks holds the promise of greater research efficiency through a more rapid recognition of physiological tissue responses.
THE NATURE AND IMPORTANCE of endogenous protective or recovery responses following sublethal acut... more THE NATURE AND IMPORTANCE of endogenous protective or recovery responses following sublethal acute physical injury have received little attention in investigations of CNS physical injury. The symposium focused attention on this important area of trauma research. An ...
An in vitro investigation was undertaken to provide information regarding the effectiveness of me... more An in vitro investigation was undertaken to provide information regarding the effectiveness of methylprednisolone sodium succinate (MPSS) as a treatment for the primary mechanical injury of spinal cord (SC) trauma. Exposure of uninjured mouse SC cells to MPSS for 24 h caused neuronal stress when the concentration exceeded 150 micrograms/mL; neuronal death occurred at concentrations above 600 micrograms/mL. The concentration range for MPSS protection of SC neurons subjected to a defined physical injury (laser microbeam transection of a primary dendrite 100 microns from the perikaryon) was very narrow: survival in the 30 micrograms/mL group differed significantly from the untreated control group (68.5% +/- 14.1 vs. 47.1% +/- 14.1), treatment with 20 or 60 micrograms/mL MPSS did not increase survival, and treatment with 100 micrograms/mL MPSS accelerated ultrastructural deterioration and increased the likelihood of death. Enhanced survival of lesioned neurons was observed when 30 micrograms/mL MPSS was applied within 15 min of dendrotomy but not when MPSS was administered 2 h after lesioning. Multimicroelectrode plate (MMEP) studies of SC network electrical activity indicated that MPSS associated readily with neuronal membranes. This finding was consistent with the hypothesis that MPSS may protect lesioned neurons by stabilizing damaged membranes, enhancing lesion resealing, and limiting the spread of ion-mediated damage. However, comparisons of neurite die-back 24 h after dendrotomy found no significant difference between MPSS-treated and control neurons. Application of 30 or 100 micrograms/mL MPSS increased the spontaneous burst activity of SC networks grown on MMEPs, however, there was no evidence that the increased excitability at these concentrations was the result of specific actions of MPSS on GABA or NMDA synapses.
Implanting electrical devices in the nervous system to treat neural diseases is becoming very com... more Implanting electrical devices in the nervous system to treat neural diseases is becoming very common. The success of these brain-machine interfaces depends on the electrodes that come into contact with the neural tissue. Here we show that conventional tungsten and stainless steel wire electrodes can be coated with carbon nanotubes using electrochemical techniques under ambient conditions. The carbon nanotube coating enhanced both recording and electrical stimulation of neurons in culture, rats and monkeys by decreasing the electrode impedance and increasing charge transfer. Carbon nanotube-coated electrodes are expected to improve current electrophysiological techniques and to facilitate the development of long-lasting brain-machine interface devices.
Prolonged (2-6 h) cooling of monolayer cultures of dissociated murine spinal cord at temperatures... more Prolonged (2-6 h) cooling of monolayer cultures of dissociated murine spinal cord at temperatures below 17 degrees C caused pronounced swelling of neuronal perikarya and dendrites. The numbers of swollen neurons in a culture increased as the temperature was reduced, and at 7 degrees C-10 degrees C all of the neurons were swollen. On rewarming the cultures to 37 degrees C, the majority of the swollen neurons died (up to 74% at 10 degrees C). Glial cells were not affected. Addition of the NMDA antagonists D-2-amino-5-phosphonovalerate (DAPV, 100 microM), ketamine (100 microM), and dibenzocyclohepteneimine (MK801, 10 microM) to spinal cord cultures before lowering the temperature to 10 degrees C minimized the dendrosomatic swelling and reduced neuronal mortality from 74% to 10%. These data show a surprising sensitivity of some neurons to nonfreezing low temperatures and suggest direct involvement of the NMDA receptor in hypothermia-related neuronal death.
Bodian and Nissl procedures were combined to stain dissociated mouse spinal cord cells cultured o... more Bodian and Nissl procedures were combined to stain dissociated mouse spinal cord cells cultured on coverslips. The Bodian technique stains fine neuronal processes in great detail as well as an intracellular fibrillar network concentrated around the nucleus and in proximal neurites. The Nissl stain clearly delimits neuronal cytoplasm in somata and in large dendrites. A combination of these techniques allows the simultaneous depiction of neuronal perikarya and all afferent and efferent processes. Costaining with little background staining by either procedure suggests high specificity for neurons. This procedure could be exploited for routine network analysis of cultured neurons.
The Journal of neuroscience : the official journal of the Society for Neuroscience, 1986
Laser-deinsulated, printed-circuit electrodes integrated into the floor of culture chambers have ... more Laser-deinsulated, printed-circuit electrodes integrated into the floor of culture chambers have been used to monitor the spontaneous activity of mouse spinal monolayer cell cultures. This technique has allowed a multisite analysis of activity over long periods of time in closed chambers. In 63 cultures investigated 3-5 weeks after seeding, 89% included single- or multiunit bursting. Based on a subset of 40 cultures in which all electrodes were sequentially scanned, bursting was found on 41% of the active electrodes (approximately 38% of all units monitored). A total of 35% of the electrodes monitoring spontaneous bursting activity revealed rhythmic sequences that were usually coupled among multiple electrodes. Although most of this coupling was in-phase, three out of 40 cultures exhibited antiphasic bursting. In all cases where coupling was observed, each electrode monitored different burst compositions, demonstrating that the activity was generated by different units. Some rhythmi...
Cultured spinal cord networks grown on microelectrode arrays display complex patterns of spontane... more Cultured spinal cord networks grown on microelectrode arrays display complex patterns of spontaneous burst and spike activity. During disinhibition with bicuculline and strychnine, synchronized burst patterns routinely emerge. However, the variability of both intra- and interculture burst periods and durations are typically large under these conditions. As a further step in simplification of synaptic interactions, we blocked excitatory AMPA synapses with 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzoquinoxaline-7-sulphonamide (NBQX), resulting in network activity mediated through the N-methyl-D-aspartate (NMDA) receptor (NMDA(ONLY)). This activity was APV sensitive. The oscillation under NMDA(ONLY) conditions at 37 degrees C was characterized by a period of 2.9 +/- 0.3 s (16 separate cultures). More than 98% of all neurons recorded participated in this highly rhythmic activity. The temporal coefficients of variation, reflecting the rhythmic nature of the oscillation, were 3.7, 4.7, and 4...
The Journal of neuroscience : the official journal of the Society for Neuroscience, 1983
The exposure of neuronal and glial cell processes to a large number (up to 300) of 12-nsec laser ... more The exposure of neuronal and glial cell processes to a large number (up to 300) of 12-nsec laser pulses at a wavelength of 337 nm and energy densities below the threshold for nonlinear absorption results in a gradual, gentle process transection in the laser focus. Within 10 to 20 sec after cessation of firing, the process pinches in the target area. During this time, mitochondria become swollen and bleached, the plasma membrane develops an obvious tautness, microtubules disappear, and organelles accumulate to either side of the process constriction. Depending on the irradiation parameters, a local pinching may proceed to a transection in about 30 sec or it may reverse to yield a normal-appearing process in approximately 5 min. Severe process pinching is accompanied by a sudden depolarization that may last for 2 to 5 min and is usually followed by a repolarization to the original resting potential even if the process has transected. Spiral retraction of cut processes and cytoplasmic ...
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