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A C Dolphin

Publications and source records attributed to A C Dolphin.

At least 91 records · Page 5Linked to original sources
PubMed1990

A comparison of the effect of calcium channel ligands and GABAB agonists and antagonists on transmitter release and somatic calcium channel currents in cultured neurons.

Glutamate release has been examined from cultured cerebellar granule neurons in the rat using the technique of prelabelling the releasable pool of glutamate with [3H]glutamine. Glutamate release was stimulated in control neurons by 2-min incubation with 50 mM K+, or in neurons continuously depolarized in Ca2(+)-free 50 mM K+ medium, by 2-min incubation with medium containing 5 mM Ca2+. The ability of the Ca2(+)-channel agonist (+)-202-791 to increase the stimulated release of [3H]glutamate was approximately doubled in the depolarized condition. The antagonist enantiomer (-)-202-791 produced a small inhibition of K(+)-stimulated release, whereas (-)-202-791 completely inhibited Ca2(+)-stimulated release from depolarized neurons at concentrations greater than 10 nM. (-)-Baclofen (100 microM) inhibited transmitter release similarly (25-30%) under the two conditions. Calcium-channel currents were recorded from cultured dorsal root ganglion neurons under control conditions at a holding potential of -80 mV, or in neurons depolarized to -30 mV. (-)-202-791 produced a greater effect at -30 than at -80 mV although even at -30 mV the inhibition was slow in onset and incomplete. (-)-Baclofen (100 microM) inhibited the amplitude of the calcium-channel current at both holding potentials by 30-50%, although it did not clearly slow activation of the current at the depolarized holding potential. The GABAB receptors associated with inhibition of glutamate release and of calcium-channel currents were both markedly blocked by phaclofen but not by 2-OH-saclofen. These findings suggest that the GABAB receptor associated with inhibitory modulation of transmitter release, and that associated with inhibition of calcium-channel currents show pharmacological similarities, and are able to exert their action even at levels of steady depolarization at which most N-type channels should be inactivated.

Animals↗
PubMed1990

Activation of calcium channel currents in rat sensory neurons by large depolarizations: effect of Guanine nucleotides and (-)-baclofen.

Calcium channel currents have been recorded from cultured rat sensory neurons at clamp potentials of between -30 and +120 mV. At large depolarizing potentials between +50 and +120 mV, the current was outward. This outward current was shown to be largely due to ions passing through calcium channels, because it was substantially although generally incompletely blocked by Cd2+ (1 mM) and omega-conotoxin (1 microM). Internal GTP-gamma-S (100 microM) and to a lesser extent GTP (1 mM) reduced the amplitude and slowed the activation of the outward, as well as the inward calcium channel current. Baclofen (100 microM) reversibly inhibited both the inward and outward currents. These results suggest that the effect of baclofen and G protein activation on calcium channel currents is not due to a shift in the voltage-dependence of channel availability.

Journal Article↗
PubMed1990

Voltage-dependent modulation of rat sensory neurone calcium channel currents by G protein activation: effect of a dihydropyridine antagonist.

The ability of a depolarizing prepulse to increase the rate of activation of IBa has been examined in cultured sensory neurones of the rat. Both in control neurones and in the presence internally of the guanine nucleotide analogue, guanosine 5'-O-3-thiotriphosphate (GTP gamma S) which markedly slows the rate of activation of IBa and reduces its amplitude, a depolarizing prepulse increased the rate of activation of IBa, but did not increase its amplitude measured at the end of the 100 ms voltage step. The calcium channel antagonist (-)-202-791, which we have previously shown to increase the amplitude of IBa in the presence of GTP gamma S, did not occlude the response to a depolarizing prepulse, suggesting that the mechanism of action of (-)-202-791 is not to disrupt the interaction of the channels with activated G proteins.

Animals↗
PubMed1990

Direct and indirect modulation of neuronal calcium currents by G-protein activation.

Evidence is presented for modulation of the various components of neuronal Ca2+ currents by G-proteins and by gamma-aminobutyric acid B (GABAB) receptor activation. The mechanism of these interactions and the possible involvement of second messengers is discussed. The role of inhibition of Ca2+ channels in presynaptic inhibition of neurotransmitter release is examined.

Animals↗
PubMed1990

Inhibition of neuronal Ca2+ channel currents by the funnel web spider toxin omega-Aga-IA.

Ca2+ channel currents were recorded from cultured rat dorsal root ganglion neurons and cerebellar granule cells using the whole-cell recording variant of the patch clamp technique. omega-Aga-IA, a toxin purified from the venom of the American funnel web spider, Agelenopsis aperta, markedly inhibited high threshold barium currents (lBa) when applied at 10 nM concentration. The low threshold T-type current activated at Vc = -30 mV and the outward (Ca2+ channel) current activated at +120 mV were significantly less sensitive to omega-Aga-IA, omega-Conotoxin GVIA (1 microM) inhibited IBa irreversibly. In contrast, the action of omega-Aga-IA was partially reversed 5 min after its removal. The voltage-activated calcium current (ICa) was inhibited by omega-Aga-IA in a manner different from IBa. ICa measured at the end of a 100-msec voltage step command was reduced to a greater extent than the peak current. The residual ICa following application of omega-Aga-IA was a fast transient current. omega-Aga-IA did not inhibit voltage-activated sodium currents from dorsal root ganglion neurons in the absence of tetrodotoxin. omega-Aga-IA abolished the dihydropyridine (+)-202-791-sensitive L-type current component of IBa. We conclude that omega-Aga-IA is a very potent inhibitor of neuronal voltage-activated Ca2+ channel currents and that it may prove to be a useful tool in the characterization and isolation of Ca2+ channels.

Agatoxins↗
PubMed1989

Bradykinin enhances excitability in cultured rat sensory neurones by a GTP-dependent mechanism.

The excitatory action of bradykinin (Bk; 0.1-1.0 microM) on cultured rat dorsal root ganglion neurones (DRGs) was studied using the whole cell clamp technique. In a subpopulation of DRGs, a 1 s depolarising voltage pulse from -70 to +20 mV evoked more than one inward current. In these neurones, local application of Bk increased the inward current frequency from 7.0 +/- 0.7 s-1 to 14.9 +/- 1.0 s-1 (mean +/- S.E.M., n = 53). Intracellular application of the GTP analogue, guanosine 5'O-3-thiotriphosphate (GTP gamma S) mimicked this excitatory action of Bk: the frequency of inward currents increased from 5.0 +/- 0.8 s-1, 30 s after the start of recording to 6.9 +/- 1.1 s-1 at 5 min to a maximum of 18.5 +/- 2.2 s-1 at 15 min (n = 16). In control cells, the frequency decreased from 4.6 +/- 0.8 s-1 to 2.5 +/- 0.5 s-1 at 5 min (n = 12). Bk also increased excitability in 4/11 Herpes Simplex Virus I (HSV-I)-infected DRGs. Thus, we demonstrate an excitatory action of Bk in DRGs, which may involve G-protein activation.

Action Potentials↗
PubMed1989

An investigation into the mechanisms of inhibition of calcium channel currents in cultured sensory neurones of the rat by guanine nucleotide analogues and (-)-baclofen.

1. The mechanism of inhibition of calcium channel currents by the guanine nucleotide analogue guanosine 5'-O-3 thiotriphosphate (GTP-gamma-S) and by the GABAB agonist (-)-baclofen has been studied in cultured dorsal root ganglion neurones of the rat. The inhibition by GTP-gamma-S is particularly characterized by an abolition of the transient component of calcium channel currents carried either by Ba2+ (IBa) or by Ca2+ (ICa). 2. The effect of agents increasing intracellular cyclic AMP levels has been examined. Neither internal cyclic AMP nor forskolin prevented the inhibition of IBa by baclofen. Neither forskolin nor pretreatment of cells with cholera toxin prevented the inhibition of the transient component of IBa by GTP-gamma-S. However, both these treatments increased the amplitude of the sustained IBa in the presence of GTP-gamma-S. The ATP analogue adenosine imido-diphosphate which inhibits many ATP requiring enzymes did not prevent the effect of GTP-gamma-S although it reduced the amplitude of IBa. 3. Baclofen (100 microM) produced a 22 +/- 2% increase in inositol phosphate production in 30 s, whereas the increase produced by bradykinin (1 microM) was 70 +/- 14%. However, unlike baclofen, bradykinin did not inhibit IBa or ICa in these cells. 4. The effect of protein kinase C inhibitors was examined. Polymixin B (20 microM in patch pipette) had no effect on the inhibition of IBa by baclofen or GTP-gamma-S. A higher concentration (100 microM) alone inhibited IBa and no further inhibition by baclofen was observed. Neither H7 (50 microM) nor staurosporine (100 nM), applied extracellularly, prevented the response to GTP-gamma-S. 5. The protein kinase C activator di-octanoyl glycerol (20 microM) did not inhibit IBa. Arachidonic acid (100 microM) also produced no inhibition of IBa. 6. In conclusion we have obtained no evidence that a second messenger system mediates the inhibition of calcium channel currents by GTP-gamma-S or baclofen in dorsal root ganglion neurones. These results support the hypothesis that GABAB receptors are directly coupled to calcium channels by G proteins.

Animals↗
PubMed1989

Interaction between calcium channel ligands and guanine nucleotides in cultured rat sensory and sympathetic neurones.

1. Voltage-activated Ca2+ channel currents were recorded from cultured rat dorsal root ganglion (DRG) neurones using the whole-cell clamp technique with Ba2+ as the charge carrier. 2. Inclusion of the GTP analogue guanosine 5'-O-3-thiotriphosphate (GTP-gamma-S, 500 microM) or guanylylimidodiphosphate (GMP-PNP, 500 microM) or GTP itself (1 mM) in the patch pipette solution resulted in a smaller, slowly activating Ca2+ channel current which did not inactivate during a 100 ms voltage step. This current was inhibited by CdCl2 (10-100 microM) and omega-conotoxin (1 microM). 3. Nifedipine (5 microM), (-)-(R)-201-791 (5 microM), D600 (10 microM), and diltiazem (30 microM) inhibited Ca2+ channel currents recorded from control neurones, although in some cells a biphasic response was observed, with an initial increase preceding the inhibition of the currents. In the presence of internal GTP-gamma-S, at a holding potential (VH) of -80 mV, only potentiation of the Ca2+ channel current was observed in the presence of all three Ca2+ channel ligands. Internal GMP-PNP, while less effective than GTP-gamma-S, also resulted in D600 showing an agonist response. Similarly, in the presence of internal GTP (1 mM), (-)-(R)-202-791 gave a prolonged agonist response. 4. Nifedipine, whether acting as an antagonist in control cells or as an agonist in GTP-gamma-S-containing cells, induced a shift to more hyperpolarized potentials of the steady-state inactivation curves. 5. Potentiation of Ca2+ channel currents induced by D600 in GTP-gamma-S-containing cells, was not observed when the neurones were pre-treated with pertussis toxin. The presence of internal GDP-beta-S (500 microM) did not significantly alter the maximum inhibitory action of D600 compared with controls. However, 1 mM-GDP-beta-S increased the rate of onset of inhibition by (-)-(R)-202-791. 6. Depolarizing VH to -30 mV accelerated the onset of inhibition induced by the Ca2+ channel ligands in control cells. In the presence of internal GTP-gamma-S at VH -30 mV, biphasic responses were produced by all the Ca2+ channel antagonist ligands with initial stimulation for 1-2 min being followed by inhibition of the Ca2+ channel currents. 7. The agonist actions of (+)-(S)-202-791 were potentiated by the presence of internal GTP-gamma-S. 8. The expression of an agonist response to (-)-(R)-202-791 induced by internal GTP-gamma-S was also present in sympathetic neurones cultured from adult rat superior cervical ganglion (SCG).(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1988

Long-term potentiation in the dentate gyrus of the anaesthetized rat is accompanied by an increase in protein efflux into push-pull cannula perfusates.

Changes in protein content of push-pull cannula perfusates from the dentate gyrus of anaesthetized rats were analyzed before and after the induction of long-term potentiation (LTP). LTP, induced by either high-frequency stimulation of the perforant path or raising the extracellular calcium concentration, was associated with increases in the protein content of the perfusates. Tetanically induced LTP was accompanied with a large but delayed increase (apparent in the second hour after the stimulation) in protein efflux. In contrast, when LTP was induced by the elevation of extracellular calcium concentration, a smaller but more immediate increase in protein efflux was observed. When 5-D-aminophosphonovalerate was used to block the induction of LTP, no increase was observed in either case. These results indicate that LTP in the dentate gyrus is accompanied by an increase in the efflux of proteins into push-pull cannula perfusates. The possible origins of these proteins and their role in LTP are discussed.

2-Amino-5-phosphonovalerate↗
PubMed1988

The agonist effect of Bay K 8644 on neuronal calcium channel currents is promoted by G-protein activation.

Voltage-activated Ca2+ channel currents were recorded from cultured rat dorsal root ganglion neurones using the whole cell clamp technique. Inclusion of 500 microM guanosine 5'-O(3-thio)triphosphate (GTP-gamma-S) in the patch pipette solution resulted in inhibition of the transient component of the Ca2+ channel current. The slowly activating, relatively non-activating Ca2+ channel current which remained in the presence of intracellular GTP-gamma-S was potentiated by the 1,4-dihydropyridine Ca2+ channel agonist Bay K 8644 (5 microM). The percentage increase induced by Bay K 8644 was greater in cells with internal GTP-gamma-S than in control cells. Pretreating cells with pertussis toxin resulted in Bay K 8644 inhibiting rather than increasing Ca2+ channel currents. These results support the hypothesis that L-type Ca2+ channel activity underlies the sustained Ca2+ channel currents which are not inhibited by GTP-gamma-S and that Ca2+ channel ligands require activation of a pertussis toxin-sensitive G protein to exert their agonist effects.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1988

Photoactivation of intracellular guanosine triphosphate analogues reduces the amplitude and slows the kinetics of voltage-activated calcium channel currents in sensory neurones.

The influence of guanine nucleotide analogues on calcium channel currents in cultured rat dorsal root ganglion neurones has been studied using a technique in which the rate of diffusion of the analogues to their site of action is by-passed by photochemical release of the analogues within the neurones. The 1(2-nitrophenyl)ethyl P3-ester derivatives of guanosine 5'-0(3-thio)triphosphate (caged GTP-gamma-S) and 5'-guanylylimidodiphosphate (caged GMP-PNP) were synthesised and found to be completely photolysable by light, yielding free GTP-gamma-S and GMP-PNP. Calcium channel currents were recorded using the whole cell patch technique and either caged GTP-gamma-S or caged GMP-PNP (2 mM) were included in the patch pipette. Stable currents were recorded for 5-10 min, and a single pulse of 300-350 nm irradiation was directed using a liquid light guide onto the recording dish. Calcium channel currents were then recorded every 30-120 s following photochemical release of approximately 20 microM GTP-gamma-S. The peak calcium channel current was reduced by about 70% with a slow time course [t1/2 1.5 +/- 0.2 min (mean +/- SEM); n = 5]. The transient component of the peak current was usually completely abolished, whereas the sustained current measured at the end of the 100 ms depolarising pulse was less affected. Qualitatively similar effects were observed on photolysis of caged GMP-PNP.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Modulation of divalent cation-activated chloride ion currents.

1. Voltage-sensitive calcium channel currents carried by Ca2+ (ICa) or Ba2+ (IBa) were followed by tail currents carried by Cl- ions in approximately 45% of cultured dorsal root ganglion neurones. 2. Extracellular application of (-)-baclofen (100 microM) inhibited IBa and ICl(Ba). Bay K 8644 (5 microM) potentiated both currents. 3. Intracellular GTP-gamma-S increased the proportion of neurones in which ICl(Ba) was recorded. In addition, the activation by GTP-gamma-S of a pertussis toxin-sensitive GTP binding (G)-protein resulted in a steady increase in the Cl- tail current with time, despite a concurrent reduction in IBa. 4. Extracellular application of 10mM caffeine selectively reduced ICl(Ba) without significant change in IBa. When Ca2+ was the charge carrier, caffeine had little effect on ICl(Ca), and increased the inactivation of ICa. 5. We conclude that, in addition to being regulated by divalent cation entry through Ca2+ channels, the Cl- current is also regulated by G-protein activation. The mechanism of activation of ICl(Ba) may involve Ca2+ release from intracellular stores.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗