Sensitivity to Adenosine of Hippocampal Pyramidal Neurones in Calcium-Free Medium

Hosseinzadeh, Hossein (1994) Sensitivity to Adenosine of Hippocampal Pyramidal Neurones in Calcium-Free Medium. PhD thesis, University of Glasgow.

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Biochemical studies have shown that divalent cations modulate adenosine receptor binding. In this electrophysiological study the effect of calcium on the postsynaptic sensitivity to adenosine was investigated. Extracellular recordings were made in the CA1 pyramidal cell layer of rat hippocampal slices following orthodromic stimulation of Schaffer collateral fibres in the stratum radiatum or antidromic stimulation of the alveus. Field excitatory postsynaptic potentials (fEPSPs) were recorded in stratum radiatum. In antidromic stimulation experiments, CaCl2 was omitted (calcium-free medium) or reduced to 0.24 mM (low calcium medium) and in some experiments MgSO4 was increased to 2 mM or 4 mM. Adenosine and baclofen induced a concentration- dependent reduction in the amplitude of orthodromic potentials with maximum effects at 20 and 5 muM , respectively. In nominally Ca2+ free medium, multiple population spikes were obtained in response to antidromic stimulation. Adenosine had little effect on reducing the secondary spike amplitude. Kynurenic acid, an excitatory amino acid antagonist, at high concentration had no effect on secondary spikes in calcium-free or low calcium medium. 2-Chloroadenosine (1-500 muM) and R-PIA (50 muM) , which are not substrates for either the nucleoside transporters or adenosine deaminase were inactive in the absence of calcium. S-(2-Hydroxy-5 nitrobenzy1)-6-thioinosine , an adenosine uptake blocker, at a concentration of 100 M-M had no effect on secondary potential size and did not restore adenosine sensitivity in calcium free medium. Sensitivity to adenosine in calcium-free medium was restored by 240 muM calcium medium or by raising magnesium (0.8-2.8 mM) . Raising the divalent cations concentration increased the inhibitory effect of adenosine and desensitisation was seen. Thapsigargin, which discharges intracellular calcium stores, at 1 muM had no significant effect on the bursts and did not change the effect of 0.5 mM adenosine in calcium free medium. Unlike adenosine, baclofen concentration-dependently reduced the secondary spike size in calcium free medium and at maximum effect (0.5 mM) or above no sign of recovery was observed during maintained superfusion for up to 45 minutes. The activity of adenosine was restored in the presence of the stabilizer agents procaine or carbamazepine, known inhibitors of sodium channels. The GABAg agonist baclofen did not restore sensitivity to adenosine. The xanthines theophylline and eyelopentyltheophylline , the latter of which is selective for Aj purine receptors, depressed the excitability of hippocampal pyramidal neurones in calcium-free media. Chelating residual calcium with EGTA reduced excitability which was additive with the xanthine effect, while 100 muM calcium depressed the response to theophylline. The inhibition by xanthines was prevented by adenosine, which had no effect by itself, but was not reproduced or modified by adenosine deaminase. The xanthine effects were also prevented by baclofen and carbamazepine. Tolbutamide 1 mM blocked the inhibitory effect of adenosine on the size of orthodromic population spikes but had no effect on the inhibitory action of adenosine on field EPSPs. Tolbutamide did not change the inhibitory action of baclofen on the population spikes or antidromic secondary spikes induced in calcium-free media with high magnesium but dramatically blocked the effect of adenosine in the calcium-free media. Levcromakalim 100 muM potentiated the inhibitory effect of adenosine, but not baclofen, on orthodromic population spikes. The results of this study showed that the mechanisms of postsynaptic activity of adenosine and baclofen are different and at postsynaptic, but not presynaptic, sites adenosine may activate an ATP-(tolbutamide or levcromakalim) sensitive potassium channel. Loss of postsynaptic sensitivity to adenosine in calcium-free solution may result from increased sodium conductances. A common feature of adenosine, baclofen and carbamazepine which may account for their antagonism of the xanthines is the blockade of calcium fluxes. It is proposed that in the presence of low external concentrations of calcium xanthines can reduce excitability by promoting the mobilisation and trans-membrane movement of residual calcium in the medium or neuronal membranes.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: Trevor Stone
Keywords: Pharmacology, Neurosciences
Date of Award: 1994
Depositing User: Enlighten Team
Unique ID: glathesis:1994-74913
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 27 Sep 2019 15:13
Last Modified: 27 Sep 2019 15:13

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