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N L Harrison

Publications and source records attributed to N L Harrison.

At least 55 records · Page 3Linked to original sources
PubMed1995

On the mechanism of modulation of transient outward current in cultured rat hippocampal neurons by di- and trivalent cations.

1. The mechanisms of Zn2+ modulation of transient outward current (TOC) were studied in cultured rat hippocampal neurons, using the voltage-clamp technique. In the presence of micromolar concentrations of external Zn2+, the voltage dependence of activation and inactivation was shifted to more positive membrane potentials. The gating of TOC was unaltered by internal application of Zn2+. The effect of Zn2+ were not mimicked by external Ca2+, except at very high concentrations (> 10 mM). 2. The modulatory effects of external Zn2+ on TOC gating were not reproduced, antagonized, nor enhanced by lowering external ionic strength, indicating that modulation by Zn2+ does not occur via screening of bulk surface negative charge. 3. A range of other divalent and trivalent metal ions also was studied, and several were found to modulate the transient outward current when added to the extracellular medium. In particular, Pb2+, La3+, and Gd3+ were potent modulators, showing activity in the low micromolar range. Other metal ions were weaker modulators (e.g., Cd2+) or were without activity at the concentrations tested (Fe3+, Cu2+, Ni2+). 4. The same range of ions also was tested on the delayed rectifier K+ current in cultured rat hippocampal neurons. None of the ions studied had significant effects on delayed rectifier gating, although high (> or = 100 microM) concentrations of Pb2+ and La3+ reduced maximal current amplitude, suggesting the possibility of channel block.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Modulation of the GABAA receptor by propofol is independent of the gamma subunit.

Many anxiolytics, anticonvulsants and general anesthetics modulate gamma-aminobutyric acid type A (GABAA) receptors. The anxiolytic benzodiazepines potentiate the actions of GABA, and this only at GABAA receptors with gamma subunits. The general anesthetics both potentiate GABA and activate GABAA receptors directly, but their binding sites on the receptor are poorly defined. We examined whether the gamma 2 subunit was required for the modulation of GABAA receptors by the general anesthetic 2,6-diisopropylphenol (propofol). Using the patch-clamp technique, we recorded membrane currents from HEK293 cells transfected with human alpha 2, beta 1 and gamma 2 cDNAs and with alpha 2 and beta 1 cDNAs alone. Both forms of the receptor were activated by GABA and by propofol at low concentrations. At maximal doses, propofol was considerably less effective than GABA as an activator of alpha 2 beta 1 GABAA receptors, but it had an efficacy similar to that of GABA as an activator of alpha 2 beta 1 gamma 2s receptors. In addition to activating currents directly, propofol potentiated currents elicited by GABA recorded from cells expressing either subunit combination. We conclude that the gamma 2 subunit is not a prerequisite for activation of GABAA receptors by propofol or for its potentiation of GABA-activated currents. However, the subunit may contribute to the efficacy of propofol as a GABAA receptor activator.

Cells, Cultured↗
PubMed1994

Biphasic modulation of the strychnine-sensitive glycine receptor by Zn2+.

The effects of extracellular applications of Zn2+ ions on the strychnine-sensitive glycine receptor were studied in cultured rat spinal cord neurons and with recombinant glycine receptors expressed in human embryonic kidney 293 cells. Nanomolar concentrations of Zn2+ enhanced the chloride ion current in response to brief applications of 100 microM glycine. The enhancement of glycine responses increased from 20 nM to 1 microM Zn2+. Higher concentrations of Zn2+ caused a reversal of the potentiation, followed by progressive inhibition of the glycine response up to approximately 20-50 microM Zn2+. The biphasic modulation by Zn2+ appeared essentially identical in native and recombinant glycine receptors. Biphasic Zn2+ modulation was observed both with picrotoxin-insensitive heteromeric (alpha 2/beta) receptors and with picrotoxin-sensitive homomeric receptors consisting only of alpha 2 subunits. This suggests that the alpha subunit alone is sufficient for formation of two distinct Zn2+ binding sites on the glycine receptor. The demonstration of Zn2+ modulation of the strychnine-sensitive glycine receptor is of potential physiological importance, in view of the likely range of subsynaptic Zn2+ concentrations to which the receptor is exposed.

Animals↗
PubMed1994

Potentiation of gamma-aminobutyric acidA receptor Cl- current correlates with in vivo anesthetic potency.

Ten general anesthetics of varying structure and potency were investigated for possible modulatory effects on gamma-aminobutyric acid, (GABAA) receptors, using the voltage clamp technique. All 10 anesthetics studied were observed to prolong the duration of responses to exogenously applied GABA recorded in cultured rat hippocampal neurons. These modulatory effects of the anesthetics occurred at pharmacologically relevant concentrations. An excellent correlation exists between drug potency as modulators of the GABAA receptor and anesthetic potency in vivo. These data suggest an alternative interpretation of the historical association between anesthetic potency and lipophilicity. It is proposed that hydrophobic binding sites on ligand-gated ion channel proteins, such as the GABAA receptors, constitute a molecular target site for many general anesthetics.

Anesthetics, General↗
PubMed1993

Calcium homeostasis in rat septal neurons in tissue culture.

Septal neurons from embryonic rats were grown in tissue culture. Microfluorimetric and electrophysiological techniques were used to study Ca2+ homeostasis in these neurons. The estimated basal intracellular free ionized calcium concentration ([Ca2+]i) in the neurons was low (50-100 nM). Depolarization of the neurons with 50 mM K+ resulted in rapid elevation of [Ca2+]i to 500-1,000 nM showing recovery to baseline [Ca2+]i over several minutes. The increases in [Ca2+]i caused by K+ depolarization were completely abolished by the removal of extracellular Ca2+, and were reduced by approximately 80% by the 'L-type' Ca2+ channel blocker, nimodipine (1 microM). [Ca2+]i was also increased by the excitatory amino acid L-glutamate, quisqualate, AMPA and kainate. Responses to AMPA and kainate were blocked by CNQX and DNQX. In the absence of extracellular Mg2+, large fluctuations in [Ca2+]i were observed that were blocked by removal of extracellular Ca2+, by tetrodotoxin (TTX), or by antagonists of N-methyl D-aspartate (NMDA) such as 2-amino 5-phosphonovalerate (APV). In zero Mg2+ and TTX, NMDA caused dose-dependent increases in [Ca2+]i that were blocked by APV. Caffeine (10 mM) caused transient increases in [Ca2+]i in the absence of extracellular Ca2+, which were prevented by thapsigargin, suggesting the existence of caffeine-sensitive ATP-dependent intracellular Ca2+ stores. Thapsigargin (2 microM) had little effect on [Ca2+]i, or on the recovery from K+ depolarization. Removal of extracellular Na+ had little effect on basal [Ca2+]i or on responses to high K+, suggesting that Na+/Ca2+ exchange mechanisms do not play a significant role in the short-term control of [Ca2+]i in septal neurons. The mitochondrial uncoupler, CCCP, caused a slowly developing increase in basal [Ca2+]i; however, [Ca2+]i recovered as normal from high K+ stimulation in the presence of CCCP, which suggests that the mitochondria are not involved in the rapid buffering of moderate increases in [Ca2+]i. In simultaneous electrophysiological and microfluorimetric recordings, the increase in [Ca2+]i associated with action potential activity was measured. The amplitude of the [Ca2+]i increase induced by a train of action potentials increased with the duration of the train, and with the frequency of firing, over a range of frequencies between 5 and 200 Hz. Recovery of [Ca2+]i from the modest Ca2+ loads imposed on the neuron by action potential trains follows a simple exponential decay (tau = 3-5 s).

Amino Acids↗
PubMed1993

Effects of volatile anesthetics on the kinetics of inhibitory postsynaptic currents in cultured rat hippocampal neurons.

1. The effects of the volatile anesthetics enflurane, halothane, and isoflurane on gamma-aminobutyric acid (GABA) receptor-mediated inhibitory postsynaptic currents (IPSCs) were studied in cultured rat hippocampal neurons. The experimental concentrations of anesthetics were measured directly using gas chromatography. All three anesthetics increased the overall duration of IPSCs, measured as the time to half-decay (T1/2). Clinically effective concentrations of anesthetics [between 0.5 and 1.5 times MAC (minimum alveolar concentration)] produced between 100 and 400% increases in T1/2. These effects were fully reversible, and did not involve alterations in the reversal potential for the IPSC (EIPSC). 2. The decay of the IPSC was fitted as a sum of two exponential functions, yielding a fast component (tau fast = 20 ms), and a slow component (tau slow = 77 ms), such that the fast component accounted for 79% of the IPSC amplitude and 52% of the total charge transfer. All three anesthetics produced concentration-related increases in the amplitude and charge transfer of the slow component, while simultaneously decreasing the amplitude and charge transfer of the fast component. Thus T1/2 approximated tau fast under control conditions, but approximated tau slow in the presence of the anesthetics. 3. Varying the calcium chelating agents in the recording pipettes had no effect on the quality or magnitude of alterations in IPSC kinetics produced by halothane, suggesting that variations in intracellular calcium levels are not required for the effect of halothane on the time course of the IPSC. 4. The (+)-stereoisomer of isoflurane produced greater increases in the duration of the IPSC than the (-)-isomer when applied at approximately equal concentrations, suggesting that there is a structurally selective site of interaction for isoflurane that modulates the GABAA receptor. 5. These results suggest that the previously shown abilities of volatile anesthetics to potentiate responses to exogenously applied GABA and to prolong the duration of GABA-mediated synaptic inhibition may be due to an alteration in the gating kinetics of the GABAA receptor/channel complex. Prolongation of synaptic inhibition in the CNS is consistent with the physiological effects that accompany anesthesia and may contribute to the mechanism of anesthetic action.

Anesthetics↗
PubMed1993

Positive modulation of human gamma-aminobutyric acid type A and glycine receptors by the inhalation anesthetic isoflurane.

The interactions of the inhalation anesthetic agent isoflurane with ligand-gated chloride channels were studied using transient expression of recombinant human receptors in a mammalian cell line. Isoflurane enhanced gamma-aminobutyric acid (GABA)-activated chloride currents in cells that expressed heteromeric GABAA receptors consisting of combinations of alpha 1 or alpha 2, beta 1, and gamma 2 subunits and in cells that expressed receptors consisting of combinations of only alpha and beta subunits. Receptors consisting of alpha 2 and gamma 2 subunits were poorly expressed but were sensitive to isoflurane. Receptors consisting of beta 1 and gamma 2 subunits were not expressed. Isoflurane also enhanced glycine-activated chloride currents through homomeric alpha glycine receptors but did not enhance GABA currents in cells expressing homomeric rho 1 receptors. These results show that not all ligand-gated chloride channel receptors are sensitive to isoflurane and, therefore, that the anesthetic interacts with specific structural determinants of these ion channel proteins.

Amino Acid Sequence↗
PubMed1993

Zinc modulates transient outward current gating in hippocampal neurons.

The actions of zinc ions (Zn2+) on transient outward current (TOC) were studied in cultured embryonic rat hippocampal neurons and adult guinea pig CA1 neurons acutely isolated from hippocampal slices. Zn2+ (1-1000 microM) shifted both the activation and inactivation curves for the TOC in the depolarizing direction. These effects of Zn2+ were modeled, assuming a single class of binding sites for Zn2+, yielding apparent dissociation constants for Zn2+ between 10 and 35 microM. In addition, the activation kinetics of the TOC were significantly slowed by Zn2+; deactivation was slowed at the highest concentrations of Zn2. These complex actions of Zn2+ on the TOC are likely to delay repolarization and promote spontaneous activity, resulting in action potential prolongation and hyperexcitability of hippocampal neurons.

Action Potentials↗
PubMed1993

Trichloroethanol potentiates synaptic transmission mediated by gamma-aminobutyric acidA receptors in hippocampal neurons.

We have examined the actions of trichloroethanol (TCEt), the active metabolite of the general anesthetic chloral hydrate, on responses mediated by gamma-aminobutyric acid (GABA)A receptors in response to application of exogenous GABA and activation of endogenous GABAergic transmission, by using hippocampal neurons in cell culture and in brain slices. In the presence of TCEt, Cl- current activated by exogenous GABA was both enhanced in amplitude and prolonged, leading to a net increase in total charge passing through GABAA receptor channels. Prolongation of GABA-activated current increased in magnitude in a concentration-dependent manner from 0.2 to 10 mM TCEt. Inhibitory postsynaptic currents produced at synapses between pairs of cultured GABAergic neurons or by activation of interneurons in hippocampal slices were also prolonged by TCEt, at concentrations from 0.5 to 10 mM. Application of TCEt at concentrations of 1 mM and above produced a small amplitude current which was directed outwardly at -40 mV in neurons in which methylsulfate or gluconate was the major intracellular anion and directed inwardly in neurons filled with Cl-. Our observations indicate that TCEt potentiates GABAergic transmission; presumably by potentiating the function of GABAA receptors in a manner similar to barbiturate or steroid anesthetics. This action is likely to contribute to the general anesthetic effect of TCEt which occurs after chloral hydrate administration.

Action Potentials↗
PubMed1993

Modulation of gating of cloned rat and human K+ channels by micromolar Zn2+.

The actions of zinc ions on three species of K+ channels were studied using mouse fibroblasts stably transfected with a plasmid containing both the appropriate K+ channel gene and a steroid-inducible promotor. The channels studied were rKv1.1 and hKv1.5, delayed rectifiers cloned from rat and human tissue, respectively, and hKv1.4, an inactivating human K+ channel. Zn2+ shifted the activation curves for all three K+ currents in the depolarizing direction and also shifted the steady state inactivation curve for hKv1.4 in the depolarizing direction. The effect of Zn2+ was concentration dependent between 2 and 1000 microM. As a consequence of the modulation of gating, the activation kinetics of the K+ currents were slowed by Zn2+, an effect likely to delay repolarization of the neuronal action potential. The action of Zn2+ on these diverse K+ channels suggests the existence of a common Zn2+ binding domain, the occupation of which influences the voltage sensor. The resulting modulation of gating of hKv1.4 by Zn2+ may well be of physiological significance, in view of the localization of this channel in mossy fiber nerve terminals in the hippocampus, where Zn2+ is found in abundance.

Animals↗
PubMed1992

The actions of 3-aminopropanephosphinic acid at GABAB receptors in rat hippocampus.

The actions of 3-aminopropanephosphinic acid (APPA) were examined using whole-cell patch-clamp recording in rat hippocampal slice. In recordings from neurons in subfield CA1 of slices from young (2-4 weeks) and adult (greater than 2 month) rats, APPA (0.5-50 microM) produced membrane hyperpolarization and outward current under voltage-clamp. APPA also inhibited excitatory postsynaptic potentials with an IC50 of 2.3 microM, and reduced inhibitory postsynaptic potentials at concentrations from 0.1 to 1 microM. The hyperpolarizing and synaptic depressant effects of APPA were reduced by 2-OH-saclofen an antagonist at the B-type receptor for the neurotransmitter gamma-aminobutyric acid (GABA). In this preparation APPA exhibited potencies similar to those previously reported for the GABAB receptor agonist baclofen. APPA was much less effective in inhibiting synaptic transmission measured using field potential recordings. The observations made with whole-cell patch-clamp recording indicate that in hippocampus APPA acts as a potent agonist at presynaptic GABAB receptors associated with both excitatory and inhibitory synapses, and also activates postsynaptic GABAB receptors.

Action Potentials↗
PubMed1992

Induction of giant depolarizing potentials by zinc in area CA1 of the rat hippocampus does not result from block of GABAB receptors.

The possibility that zinc (Zn2+) induces giant depolarizing potentials (GDPs) by blocking pre- and postsynaptic gamma-aminobutyric acidB (GABAB) receptors in area CA1 of rat hippocampal slices was investigated. Monosynaptic GABAA receptor-mediated fast and GABAB receptor-mediated late inhibitory postsynaptic potentials (IPSPs) were evoked in the presence of the excitatory amino acid (EAA) receptor antagonists 6,7-dinitroquinoxaline-2,3-dione (DNQX) and D,L-amino-5-phosphonovalerate (APV). Addition of Zn2+ (0.3 mM) resulted in the appearance of long-lasting GDPs which obscured monosynaptic late IPSPs. The GABAA receptor antagonist bicuculline methiodide (BMI; 30 microM) blocked fast monosynaptic IPSPs and GDPs, revealing a monosynaptic late IPSP that was prolonged in the presence of Zn2+ and blocked by the GABAB receptor antagonist CGP 35,348 (100 microM). The selective GABAB receptor agonist baclofen (10 microM) depressed monosynaptic IPSPs and population excitatory postsynaptic potentials (pEPSPs) by acting at presynaptic GABAB receptors. Depression of synaptic potentials by baclofen was unaffected by Zn2+. These results suggest that induction of GDPs in area CA1 does not result from an action of Zn2+ at GABAB receptors. We suggest instead that Zn2+ induces GDPs by inducing synchronized discharge of GABAergic interneurons.

2-Amino-5-phosphonovalerate↗
PubMed1992

Investigations into neuropeptide Y-mediated presynaptic inhibition in cultured hippocampal neurones of the rat.

1. We have examined the effects of neuropeptide Y (NPY) on synaptic transmission and [Ca2+]i signals in rat hippocampal neurones grown in culture. [Ca2+]i in individual neurones displayed frequent spontaneous fluctuations often resulting in an elevated plateau [Ca2+]i. These fluctuations were reduced by tetrodotoxin (1 microM) or combinations of the excitatory amino acid antagonists 6-cyano-7-dinitro-quinoxaline (CNQX) (10 microM) and aminophosphonovalerate (APV) (50 microM), indicating that they were the result of glutamatergic transmission occurring between hippocampal neurones. 2. [Ca2+]i fluctuations were also prevented by Ni2+ (200 microM), by the GABAB receptor agonist, baclofen (10 microM) and by NPY (100 nM) or Y2 receptor-selective NPY agonists. Following treatment of cells with pertussis toxin, NPY produced only a brief decrease in [Ca2+]i fluctuations which rapidly recovered. 3. Perfusion of hippocampal neurones with 50 mM K+ produced a large rapid increase in [Ca2+]i. This increase was slightly reduced by NPY or by a combination of CNQX and APV. The effects of CNQX/APV occluded those of NPY. NPY had no effect on Ba2+ currents measured in hippocampal neurones under whole cell voltage-clamp even in the presence of intracellular GTP-gamma-S. On the other hand, Ba2+ currents were reduced by both Cd2+ (200 microM) and baclofen (10 microM). 4. Current clamp recordings from hippocampal neurones demonstrated the occurrence of spontaneous e.p.s.ps and action potential firing which were accompanied by increases in [Ca2+]i. This spontaneous activity and the accompanying [Ca2+]i signals were prevented by application of NPY (100 nM). When hippocampal neurones were induced to fire trains of action potentials in the absence of synaptic transmission, these were accompanied by an increase in cell soma [Ca2+]j. NPY (100 nM) had no effect on these cell soma [Ca2+], signals. NPY (100 nM) also had no effect on inward currents generated in hippocampal neurones by micropipette application of glutamate (50 microM).5. Thus, NPY is able to abolish excitatory neurotransmission in hippocampal cultures through a pertussis toxin-sensitive mechanism. However, no effect of NPY on Ca2+ influx into the cell soma of these hippocampal neurones could be discerned. These results are consistent with a localized presynaptic inhibitory effect of NPY on glutamate release in hippocampal neurones in culture.

2-Amino-5-phosphonovalerate↗
PubMed1992

Enhancement of gamma-aminobutyric acid-activated Cl- currents in cultured rat hippocampal neurones by three volatile anaesthetics.

1. The effects of the volatile anaesthetics enflurane, halothane and isoflurane on gamma-aminobutyric acid (GABA)A receptor-mediated chloride currents were studied in cultured rat hippocampal neurones. Transient current responses were obtained by brief pressure application of GABA to the cell body of neurones under voltage clamp. 2. All three anaesthetics increased the peak amplitude and duration of current 2. All three anaesthetics increased the peak amplitude and duration of current responses to brief applications of GABA. These effects were fully reversible, and did not involve alterations in the reversal potential for GABA responses. 3. The experimental concentrations of anaesthetics were measured directly using gas chromatography. The enhancement of GABA currents increased with increasing anaesthetic concentration. Clinically effective concentrations of anaesthetics (between 1 and 1.5 times MAC (minimum alveolar concentration) produced significant enhancement of GABA currents. 4. These results demonstrate that the changes in the time course of synaptic inhibition reported in the presence of the volatile anaesthetics are likely to result from modification of the function of postsynaptic GABAA receptor-channel complexes. These findings also support the hypothesis that GABAA receptor complexes serve as common molecular target sites for a variety of structurally diverse anaesthetic molecules.

Animals↗
PubMed1991

Baclofen-induced disinhibition in area CA1 of rat hippocampus is resistant to extracellular Ba2+.

The mechanism of disinhibition produced by (+/-)-baclofen was studied using intracellular recording in area CA1 of rat hippocampal slices. Baclofen reversibly depressed monosynaptic IPSPs evoked by direct activation of interneurons in the presence of the excitatory amino acid receptor antagonists 6,7-dinitroquinoxaline-2,3-dione (DNQX) and D,L-2-amino-5-phosphonovalerate (APV). Ba2+ prevented baclofen-induced hyperpolarization of pyramidal neurons but not depression of monosynaptic IPSPs by baclofen. Baclofen reversibly depressed monosynaptic IPSPs when applied close to the recording site, but was ineffective when applied close to the stimulating site in stratum radiatum. These results suggest that baclofen disinhibits pyramidal neurons in area CA1 of the rat hippocampus by activating receptors on the terminals of inhibitory neurons that are coupled to a Ba(2+)-insensitive effector mechanism.

2-Amino-5-phosphonovalerate↗
PubMed1991

Evidence for mu opiate receptors on inhibitory terminals in area CA1 of rat hippocampus.

The mechanism of disinhibition produced by opioid peptides was studied using intracellular recording in area CA1 of rat hippocampal slices. The mu-selective opioid peptide [D-Ala2,N-Me-Phe4,Gly-ol5]-enkephalin (DAGO) reversibly depressed directly-activated, monosynaptic inhibitory postsynaptic potentials (IPSPs) evoked in the presence of the excitatory amino acid receptor antagonists 6,7-dinitroquinoxaline-2,3-dione (DNQX) and D,L-2-amino-5-phosphonovalerate (APV) in a naloxone-sensitive manner. Depression of monosynaptic inhibitory postsynaptic potentials (IPSPs) by DAGO was not prevented by 1-2 mM Ba2+. DAGO reversibly depressed monosynaptic IPSPs when applied locally close to the recording site, but was ineffective when applied close to the stimulating site in stratum radiatum. These results suggest that DAGO disinhibits pyramidal neurons in area CA1 of the rat hippocampus by activating mu opiate receptors located on the terminals of inhibitory neurons, and by a Ba(2+)-insensitive mechanism.

2-Amino-5-phosphonovalerate↗
PubMed1990

The actions of 2-hydroxy-saclofen at presynaptic GABAB receptors in the rat hippocampus.

The actions of 2-hydroxy-saclofen (2-OH-S), a recently developed analog of baclofen, were studied at presynaptic GABAB receptors in the rat hippocampal slice. Baclofen (0.5-20 microM) reduces the amplitude of excitatory postsynaptic potentials (EPSPs) recorded from hippocampal CA1 pyramidal neurons. In the presence of 200-500 microM 2-OH-S, the synaptic depressant action of baclofen is significantly reduced. These data show that 2-OH-S is an effective antagonist at presynaptic GABAB receptors on excitatory terminals in the hippocampus.

Animals↗