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S A Lipton

Publications and source records attributed to S A Lipton.

At least 145 records · Page 8Linked to original sources
PubMed1992

Desensitization of GABA-activated currents and channels in cultured cortical neurons.

Application of GABA to rat neocortical neurons maintained in cell culture produced a response that declined over several seconds, even in the continued presence of agonist. The decrement could be attributed to both a redistribution of Cl- and a true decline in GABA-induced membrane conductance, or desensitization. The extent and rate of desensitization were dose dependent in a manner similar to the dose dependence of the GABA-induced current, but were not related to the absolute magnitude of the current or to the charge transfer. Bicuculline slowed desensitization while diazepam enhanced the rate of desensitization, consistent with a localization of desensitization to the agonist-receptor binding site. When measured in the whole-cell recording mode, desensitization was voltage dependent, becoming much slower as the membrane was depolarized. Changes in extracellular or intracellular [Ca2+] did not appear to grossly affect the desensitization process or its voltage dependence. GABA-activated single channels, recorded in the outside-out configuration, also desensitized in the continued presence of agonist. However, desensitization differed from that seen in the same neurons in the whole-cell mode. Desensitization was considerably more rapid and did not show any voltage sensitivity. Moreover, single-channel responses often failed to recover after only a few exposures to agonist. Desensitization of GABA responses may play a role in the regulation of cortical inhibition, especially under conditions of intense excitatory and inhibitory synaptic activation.

Animals↗
PubMed1992

Open-channel block of N-methyl-D-aspartate (NMDA) responses by memantine: therapeutic advantage against NMDA receptor-mediated neurotoxicity.

Excessive activation of NMDA receptors is thought to mediate the calcium-dependent neurotoxicity associated with hypoxic-ischemic brain injury, trauma, epilepsy, and several neurodegenerative diseases. For this reason, various NMDA antagonists have been investigated for their therapeutic potential in these diseases, but heretofore none have proven to be both effective and safe. In the present study, memantine, an adamantane derivative similar to the antiviral drug amantadine, is shown to block the channels activated by NMDA receptor stimulation. From whole-cell and single-channel recording experiments, the mechanism of action of memantine is deduced to be open-channel block, similar to MK-801; however, unlike MK-801, memantine is well tolerated clinically. Compared to MK-801, memantine's safety may be related to its faster kinetics of action with rapid blocking and unblocking rates at low micromolar concentrations. Furthermore, at these levels memantine is an uncompetitive antagonist and should theoretically allow near-normal physiological NMDA activity throughout the brain even in the face of pathologically high focal concentrations of glutamate. These pharmacological properties confer upon memantine a therapeutic advantage against NMDA receptor-mediated neurotoxicity with few side effects compared with other organic NMDA open-channel blockers. Moreover, memantine is increasingly effective against escalating levels of glutamate, such as those observed during a stroke. Low micromolar concentrations of memantine, levels known to be tolerated by patients receiving the drug for the treatment of Parkinson's disease, prevent NMDA receptor-mediated neurotoxicity in cultures of rat cortical and retinal ganglion cell neurons; memantine also appears to be both safe and effective in a rat stroke model. These results suggest that memantine has considerable therapeutic potential for the myriad of clinical entities associated with NMDA receptor-mediated neurotoxicity.

Animals↗
PubMed1991

Calcium channel antagonists attenuate NMDA receptor-mediated neurotoxicity of retinal ganglion cells in culture.

Dihydropyridine calcium channel antagonists block a prolonged or 'L-type' component of voltage-dependent Ca2+ current in patch-clamp recordings of postnatal rat retinal ganglion cells. In the present study on these neurons, calcium channel antagonists were found at 500-1000 nM concentrations to attenuate the early rise in [Ca2+]i and the subsequent toxic effects of exogenous glutamate, N-methyl-D-aspartate (NMDA), or an endogenous glutamate-related compound present in the retinal cultures. Previous data have shown that the neurotoxicity engendered by these agents can also be prevented by selective NMDA antagonists. The present observations raise the possibility, at least in this preparation, that activation of both voltage-dependent calcium channels and NMDA receptor-operated channels contribute to the injurious effects triggered by molecules binding to the NMDA receptor.

Animals↗
PubMed1991

GDP beta S antagonizes whole-cell current responses to excitatory amino acids.

Guanine nucleotides have been predicted to be NMDA antagonists in tissue binding studies. Using the patch-clamp technique, we now show that the guanine nucleotide GDP beta S produces a rapidly reversible antagonism of NMDA, kainate, and, less potently, quisqualate whole-cell current responses. Furthermore, it does not appear that this antagonism is effected intracellularly. Our results suggest a novel extracellular role for guanine nucleotides apart from their traditional intracellular actions on G-proteins.

Animals↗
PubMed1991

Calcium channel antagonists and human immunodeficiency virus coat protein-mediated neuronal injury.

Recent evidence in vitro has suggested that neuronal injury observed in the acquired immunodeficiency syndrome dementia complex may depend, at least in part, on toxic effects of the human immunodeficiency virus type 1 envelope protein, gp120. This laboratory previously reported that members of the dihydropyridine class of calcium channel antagonists, nimodipine and nifedipine, greatly attenuate the rise in intracellular calcium engendered by gp120 and prevent subsequent neuronal injury. The relatively low (nanomolar) concentrations of dihydropyridines that were effective suggested that their action might be exerted at the level of the L-type of voltage-dependent calcium channels. In the present study, I tested members of the three other major classes of Ca2+ channel antagonist drugs to determine if they too could prevent neurotoxicity induced by gp120. At the maximal dose that did not cause neuronal damage in and of itself, a diphenylalkylamine piperazine derivative (flunarizine, 10 microM) was the most effective, a phenylalkylamine (verapamil, 100 microM) was possibly effective, whereas a benzothiazepine (diltiazem, 1 microM) was ineffectual in protecting rat retinal ganglion cells from gp120-induced toxicity in vitro. To explain these results, previous work has shown that the various classes of Ca2+ channel antagonists may exhibit differential potency in blocking voltage-dependent Ca2+ current in neurons, with dihydropyridines and flunarizine being the most potent at neuronal calcium channels. Moreover, these channels on mammalian central neurons are relatively insensitive to agents such as verapamil and diltiazem compared with other cell types like muscle. The low micromolar concentrations necessary for potency of flunarizine is in keeping with that predicted by binding and electrophysiological studies for block of voltage-dependent calcium channels.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Redox modulatory site of the NMDA receptor-channel complex: regulation by oxidized glutathione.

We monitored increases in both intracellular calcium concentration [( Ca2+]i) and whole-cell current responses induced by N-methyl-D-aspartate (NMDA), applied with co-agonist glycine, using fura-2 digital imaging and patch-clamp recording techniques. Extracellular application of oxidized glutathione (GSSG), but not reduced glutathione (GSH), inhibited responses mediated by activation of the NMDA subtype of glutamate receptor in cultures of rat cortical and retinal ganglion cell neurons. The NMDA responses were persistently inhibited by GSSG (500 microM to 10 mM) until introduction of a selective sulfhydryl reducing agent, dithiothreitol, which resulted in complete recovery of the responses. Exposure of the neurons to 5,5-dithio-bis-2-nitrobenzoic acid (DTNB), an efficacious oxidizing agent, also resulted in persistently smaller responses to NMDA. The addition of GSSG following exposure to DTNB, however, did not result in a further decrement in NMDA responses in our experimental paradigm. These findings suggest that a predominant action of GSSG is oxidation of vicinal thiol groups to form a peptide disulfide bond(s) comprising the redox modulatory site of the NMDA receptor-channel complex. Evidence for such regulatory sulfhydryl centers associated with the NMDA receptor has been presented previously. Moreover, the fact that DTNB produced little if any additional attenuation of the NMDA [Ca2+]i response when administered after GSSG implies that GSSG is also an efficacious oxidant at this site. GSSG displayed little or no effect on [Ca2+]i responses elicited by high extracellular K+ or by kainate, suggesting that, at least under the conditions of the present experiments, GSSG was somewhat selective for the NMDA redox modulatory site. Based on these observations, we suggest that GSSG exerts its NMDA-specific redox effects in a novel extracellular manner.

Animals↗
PubMed1991

Immunofluorescent characterization of retinal ganglion cell neurites cultured on substrates coated with antibodies against Thy-1.

Neurite outgrowth from rat retinal ganglion cells is enhanced if they are cultured on a substrate coated with antibodies against Thy-1. We show that the ganglion cell neurites that grow on antibodies against Thy-1 display immunoreactivity for microtubule-associated protein 2 and tau, cytoskeletal proteins normally localized to the somatodendritic and axonal compartments, respectively. This result suggests that these neurites are incompletely differentiated and have axonal and dendritic features. We also show that the antibodies against Thy-1 used to coat the substrate bind specifically to ganglion cell membranes within 1 day of culture.

Animals↗
PubMed1991

Cryopreservation of postnatal rat retinal ganglion cells: persistence of voltage- and ligand-gated ionic currents.

Established methods for cryopreservation of living cells were modified for freeze-storage of postnatal retinal ganglion cells from rat. Retinal cell suspensions containing fluorescently labeled ganglion cells were frozen after addition of 8% dimethyl sulfoxide and stored at -80 degrees C for up to 66 days. Viability of identified retinal ganglion cells was assessed by their ability to take up and cleave fluorescein diacetate to fluorescein. No significant difference was found in the number of living retinal ganglion cells when cells obtained from the same dissociation were counted before and after freezing (6.65 +/- 2.37 x 10(4) vs 7.05 +/- 3.67 x 10(4) retinal ganglion cells per ml, respectively; mean +/- S.D., n = 4). In culture following cryopreservation, the cells appeared morphologically normal, and developed neurites and growth cones similar to their freshly dissociated counterparts. Since very little is known about the electrophysiology and membrane properties of neurons after cryopreservation, we used the whole-cell configuration of the patch-clamp technique to study voltage- and ligand-gated conductances in cryopreserved retinal ganglion cells. The cryopreserved retinal ganglion cells studied under current-clamp maintained resting potentials of -60.9 +/- 6.6 mV (n = 10) and upon depolarization fired action potentials. During voltage-clamp in the whole-cell mode, depolarizing voltage steps activated Na(+)-(INa), Ca(2+)-(ICa), and K(+)-currents in all cells tested (n = 122). INa could be reversibly blocked by 1 microM tetrodotoxin added to the external solution. ICa was blocked by external 250 microM Cd2+ or 3 mM Co2+. In some cells, ICa consisted of both a transient and prolonged component. The outward K(+)-current consisted of Ca(2+)-dependent and -independent components. The Ca(2+)-insensitive portion of the K+ outward current was separated into four distinct components based upon pharmacological sensitivity and biophysical properties. In many cells, a rapidly inactivating current similar to the A-type K(+)-current (IA) observed in freshly cultured retinal ganglion cells was isolated by its greater sensitivity to 4-aminopyridine (5 mM) than to tetraethylammonium (20 mM). A tetraethylammonium-sensitive current with a more prolonged time course reminiscent of IK, the delayed rectifier, was also found. When the 4-aminopyridine- and tetraethylammonium-insensitive portions of the outward current were further analysed with voltage protocols, an additional slowly decaying potassium current became apparent. The inhibitory amino acids, GABA (20 microM) and glycine (100 microM), activated chloride-selective currents that were selectively blocked by bicuculline methiodide (10 microM) and strychnine (5 microM), respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylcholine↗
PubMed1991

Synergistic effects of HIV coat protein and NMDA receptor-mediated neurotoxicity.

Exposure of rat retinal cultures to HIV-1 coat protein gp120 for several minutes increases [Ca2+]i in approximately half of the ganglion cells; this effect is associated with delayed-onset neuronal injury, similar to that previously reported in NMDA receptor-mediated neurotoxicity. Here we show that NMDA antagonists can prevent both the rise in [Ca2+]i and subsequent neuronal damage engendered by 20 pM gp120. However, whole-cell patch-clamp recordings demonstrate that gp120 does not directly evoke an NMDA-like response or enhance glutamate/NMDA-activated currents. Moreover, complete protection from gp120-induced [Ca2+]i increases and neurotoxicity is afforded by incubation with glutamate-pyruvate transaminase, which breaks down endogenous glutamate as verified by HPLC. Since, under standard conditions in these cultures, neither glutamate nor a low picomolar concentration of gp120 is deleterious on its own, our results suggest that their neurotoxicity is synergistic.

2-Amino-5-phosphonovalerate↗
PubMed1991

Glutathione prevents N-methyl-D-aspartate receptor-mediated neurotoxicity.

N-methyl-D-aspartate(NMDA) antagonists afford possible therapeutic modalities for stroke, trauma, and various neurodegenerative conditions thought to be mediated by excessive stimulation of NMDA receptors in the brain. To date, however, no drug has been demonstrated to be a safe NMDA antagonist at a dosage necessary for neuroprotection. In the present study, we use an in-vitro preparation to show that glutathione is capable of attenuating neuronal damage mediated by NMDA receptor activation. Both oxidized and reduced glutathione are protective, and an extracellular mechanism of action on the NMDA receptor-channel complex is suggested.

Animals↗
PubMed1991

HIV-related neurotoxicity.

The central nervous system manifestations of AIDS were originally thought to consist solely of white matter lesions, but recent evidence has shown that a substantial degree of neuronal loss can also occur. This review presents evidence for HIV-related toxic factors that may account at least in part for this newly-recognized neuronal injury. One potential neurotoxin is the HIV-1 envelope glycoprotein gp-120 or a fragment of this molecule. This coat protein is shed by the virus and potentially released from HIV-infected immune cells. In tissue culture experiments on rodent neurons, gp120 produces an early rise in intracellular calcium concentration and, subsequently, delayed-onset neurotoxicity. In addition, HIV-infected macrophages or microglia release as yet undefined toxic factor(s) that kill rodent, chick, and human neurons in vitro. It is as yet unknown if one of these macrophage toxic factors might represent a gp120 fragment, or alternatively, if gp120, in the absence of HIV-1 infection, might be capable of activating macrophages to release these toxic factor(s). In at least some neuronal cell types, gp120-induced neurotoxicity can be prevented by antagonists of L-type voltage-dependent calcium channels or by antagonists of N-methyl-D-aspartate (NMDA, a subtype of glutamate receptor). Degradation of endogenous glutamate also protects neurons from gp120-related neuronal injury, suggesting that gp120 and glutamate are both necessary for neuronal cell death as synergistic effectors. Antagonists acting at the other types of glutamate receptors (non-NMDA antagonists) are ineffective in affording protection from gp120.(ABSTRACT TRUNCATED AT 250 WORDS)

AIDS Dementia Complex↗
PubMed1991

N-methyl-D-aspartate antagonists prevent kainate neurotoxicity in rat retinal ganglion cells in vitro.

Under defined culture conditions, exogenous glutamate (Glu), NMDA, or an endogenous Glu-related toxin is lethal to rat retinal ganglion cells; these detrimental effects are NMDA receptor mediated because specific NMDA antagonists can prevent cellular injury. In the presence of an endogenous Glu-like toxin, 125 microM kainate (KA) increases the proportion of retinal ganglion cells that die, but the toxicity (due to both KA and the endogenous toxin) is totally prevented by 2-amino-5-phosphonovalerate (APV), a specific NMDA receptor antagonist. These findings indicate that the KA-induced portion of retinal ganglion cell death also appears to be mediated via NMDA receptors. There are at least 2 possible mechanisms for this lethal effect. In addition to KA receptors, KA could directly stimulate NMDA receptors. Alternatively, KA might activate its own specific receptor, which in turn leads to a net increase in the release of an endogenous Glu-related toxin; this endogenous substance would then activate NMDA receptors. Patch-clamp electrophysiology experiments have helped to distinguish between these possibilities. Concentrations of APV that completely block the current elicited by maximal nondesensitizing doses of NMDA exert no detectable inhibition of KA-evoked currents. Hence, at the concentrations used, it appears unlikely that KA directly activates NMDA receptors in this preparation. Furthermore, the fraction of toxicity attributed to the addition of KA can be blocked by the relatively specific non-NMDA antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). This finding is consistent with the hypothesis that KA adds an increment of toxicity in this system by directly interacting with KA receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

2-Amino-5-phosphonovalerate↗
PubMed1990

Neural nicotinic acetylcholine responses in sensory neurons from postnatal rat.

The whole-cell configuration of the patch-clamp technique was used to study nicotinic acetylcholine (ACh) responses in freshly dissociated dorsal root ganglion (DRG) cells from postnatal rat. At negative holding potentials with physiological solutions in the bath and the pipette, ACh (20 microM), nicotine (5 microM) or DMPP (20 microM) activated inward currents in 51% of the cells. Average current density was higher in 1-month-old compared to newborn animals. Nicotinic agonist-induced currents were unaffected by atropine (10 microM) but reversibly blocked by hexamethonium (20 microM). Although labeling with fluorescent alpha-bungarotoxin (BGT) demonstrated the presence of toxin binding sites on DRG cells, DMPP-induced inward currents were unaffected by micromolar BGT. Neuronal bungarotoxin (100 nM), in contrast, led to a largely irreversible block of the nicotinic responses. These results show that postnatal DRG cells express functional nicotinic acetylcholine receptors (nAChR) of a neuronal type.

Animals↗
PubMed1990

Blockade of nicotinic responses in rat retinal ganglion cells by neuronal bungarotoxin.

The effects of neuronal bungarotoxin (NBT) on nicotinic acetylcholine responses recorded from rat retinal ganglion cells in culture were studied with patch electrodes. We observed that the concentration of this toxin needed to induce a total blockade of nicotinic currents varied according to the method of toxin application utilized. Rapid addition of 20 microM NBT by pressure ejection from micropipettes produced total blockade of 50-microM acetylcholine-induced currents. In contrast, when added slowly via the physiological solution which continuously superfused the cells, NBT was able to produce a complete blockade of the response at 200 nM. The IC50 determined for NBT by the superfusion method was 55 nM. Recovery from block was slow and incomplete with both drug application methods, although some differences were found. The results are discussed with reference to a scheme in which NBT binds with both low and high affinity to functional nicotinic receptors in rat retinal ganglion cells in culture.

Acetylcholine↗
PubMed1990

HIV-1 coat protein neurotoxicity prevented by calcium channel antagonists.

Coat protein gp120 from the human immunodeficiency virus type-1 (HIV-1) increased intracellular free calcium and injured rodent retinal ganglion cells and hippocampal neurons in culture. Highly purified recombinant gp120 envelope protein produced these effects in a dose-dependent fashion at picomolar concentrations. Immunoprecipitation with antibody to gp120, but not with control immunoglobulin-containing serum, depleted solutions of the viral envelope protein and also prevented both the rise in intracellular calcium and neuronal toxicity. The gp120-induced increase in intracellular calcium was abrogated by transiently lowering extracellular calcium or by adding the dihydropyridine calcium channel antagonist nimodipine (100 nM). Calcium channel antagonists also prevented gp120-induced neuronal injury. In addition, intracellular stores appeared to contribute substantially to the increase in calcium elicited by gp120. Since increases in intracellular calcium have been associated with neurotoxicity, it is possible that an injurious effect of gp120 on neurons might be related to this mechanism and that treatment with calcium channel antagonists may prove useful in mitigating HIV-1-related neuronal injury.

Animals↗
PubMed1990

Redox modulation of NMDA receptor-mediated toxicity in mammalian central neurons.

Acute neurological injury from hypoxia-ischemia, hypoglycemia, and trauma is thought to be predominantly mediated by activation of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor in the brain and the subsequent influx of calcium ions through receptor-operated channels. Several chronic degenerative diseases, such as Huntington's disease and the amyotrophic lateral sclerosis-Parkinsonism-dementia complex found on Guam, may share a similar pathogenesis due to a glutamate-like toxin. This laboratory recently reported that exposure to a reducing agent, such as dithiothreitol (DTT), selectively increases ionic current flow through NMDA-activated channels in several types of central neurons; conversely, oxidizing agents reverse this effect. To investigate the novel influence of redox modulation on NMDA neurotoxicity, in the present in vitro study we monitored survival of an identified central neuron, the retinal ganglion cell, approximately 24 h after a brief exposure to DTT. To determine the degree of killing specifically related to activation of the NMDA receptor, 2-amino-5-phosphonovalerate (APV, a selective NMDA antagonist) was added to sibling cultures. APV-preventable, glutamate-induced death was increased 70 +/- 9% with DTT treatment. This effect was totally blocked by the concomitant addition of an oxidizing agent, 5,5-dithiobis-2-nitrobenzoic acid (DTNB). These findings suggest that the enhanced killing following chemical reduction with DTT is mediated at the NMDA receptor site, and that the redox state of the NMDA receptor is crucial for the survival of neurons facing glutamate-related injury.(ABSTRACT TRUNCATED AT 250 WORDS)

2-Amino-5-phosphonovalerate↗
PubMed1990

VIP-mediated increase in cAMP prevents tetrodotoxin-induced retinal ganglion cell death in vitro.

Afferent influences on natural cell death were modeled in retinal cultures derived from neonatal rats. Tetrodotoxin (TTX) blockade of electrical activity produced a significant reduction in surviving retinal ganglion cell (RGC) neurons during a critical period of development, similar in magnitude to the reduction observed during natural cell death in the intact retina at a similar developmental stage. The addition of vasoactive intestinal peptide (VIP) protected the RGCs from the lethal action of TTX. This effect was specific, since the related peptides PHI-27 and secretin produced no significant increase in RGC survival. Radioimmunoassay of cyclic nucleotides showed that TTX decreased culture levels of cAMP and that this trend was reversed by VIP. Decreases in RGC survival associated with TTX electrical blockade were prevented by 8-bromo:cAMP or forskolin. Furthermore, VIP10-28, the C-terminal fragment that inhibits VIP stimulation of adenylate cyclase, reduced the number of surviving RGCs. Thus, our results suggest that VIP, acting by increasing cAMP, has a neurotrophic effect on electrically blocked RGCs and may be an endogenous factor modulating normal cell death in the retina.

8-Bromo Cyclic Adenosine Monophosphate↗