Interaction between adenosine A₁ and A₂ receptor-mediated responses in the rat hippocampus in vitro

Abstract

Previous work has been carried out on the effects of adenosine on transmitter release and on the excitability of postsynaptic neurones, but little is known about the effects of adenosine on the coupling between the two. In this study, we examine the effects of specific adenosine receptor agonists and antagonists on the population excitatory postsynaptic potential (population EPSP) slope, the population spike amplitude, and the relationship between the two (E–S coupling) in the CA1 area of rat hippocampus. Activation of adenosine A₁ receptors by adenosine or the selective agonist N⁶-cyclopentyladenosine resulted in a decrease of the population spike amplitude by a greater extent than could be accounted for by the decrease in population EPSP slope, resulting in a dissociation in the E–S relationship, reflected as a right-shift in the E–S curve. Activation of adenosine A_2A receptors by the selective agonist 2-p-(2-carboxyethy)phenethylamino-5′-N-ethylcarboxamidoadenosine (CGS 21680), or blockade by antagonists ZM 241385 and CP 66713 had no effect on evoked responses. However, when both adenosine A₁ and A_2A receptors were activated at the same time, a significant attenuation of the inhibitory effects of N⁶-cyclopentyladenosine on population spike amplitude was observed, resulting in a left-shift in the E–S curve. Intracellular recording indicated that N⁶-cyclopentyladenosine raised the threshold for spike induction by pulses of depolarising current, even at a concentration which did not produce hyperpolarisation of the neurone. At 30 nM, CGS 21680 prevented this effect of N⁶-cyclopentyladenosine, and this apparent antagonism was prevented by the A_2A receptor antagonist ZM 241385. The results show that adenosine A₁ receptors change the coupling between presynaptic transmitter release and postsynaptic cell firing, and that this effect is attenuated by A_2A receptor activation.

Introduction

Adenosine is a neuromodulator with both presynaptic and postsynaptic effects in the mammalian central nervous system (CNS). It has been suggested that these effects are mediated by different mechanisms in view of, for example, their different sensitivities to agents such as pertussis toxin (Fredholm et al., 1989; Hasuo et al., 1992; Thompson et al., 1992). Presynaptically, adenosine has been shown to inhibit the release of neurotransmitters such as acetylcholine (Spignoli et al., 1984; Cunha et al., 1994b), dopamine (Michaelis et al., 1979; Zetterstrom and Fillenz, 1990), serotonin (Feuerstein et al., 1985), and glutamate (Fastbom and Fredholm, 1985). The mechanism of adenosine's presynaptic effects is not fully clear but it has been suggested that it causes an increase in potassium conductance which could hyperpolarise the axon terminal and so prevent transmitter release (Thompson et al., 1992). Supporting this, it has been shown that K⁺ channel blockers such as 4-aminopyridine can reduce the effects of adenosine (Stone, 1981). An indirect decrease of Ca²⁺ currents or intracellular Ca²⁺ levels by this adenosine activated potassium conductance has also been suggested as a mechanism of presynaptic inhibition (Dunwiddie and Haas, 1985; Michaelis et al., 1988). Alternatively, adenosine may exert its presynaptic effects by inhibiting Ca²⁺ influx directly (Dolphin et al., 1986; Schubert et al., 1987).

Postsynaptically, the mechanism of action of adenosine is clearer. Several groups have suggested that adenosine causes an increase in outward potassium conductance leading to a hyperpolarisation of the postsynaptic membrane (Okada and Ozawa, 1980; Segal, 1982; Trussel and Jackson, 1985). This potassium conductance has been shown to be both Ca²⁺ and voltage insensitive in CA1 neurones (Gerber et al., 1989).

Earlier studies have suggested that the inhibitory presynaptic actions and postsynaptic hyperpolarising actions of adenosine are mediated by A₁ receptors (Dunwiddie and Fredholm, 1989; Alzheimer et al., 1991; Ameri and Jurna, 1991; Lambert and Teyler, 1991; Hasuo et al., 1992) which are present in high density in the hippocampus, especially in the CA1 region (Fastbom et al., 1987). Electrophysiological (Sebastião and Ribeiro, 1992) and binding (Jarvis et al., 1989; Cunha et al., 1994a) studies have shown that adenosine A_2A receptors are also present in the hippocampus but conflicting reports exist as to the functional importance of these receptors and their ability to interact with adenosine A₁ receptors. Cunha et al. (1994a) describe an attenuation of the inhibitory effects on population spike amplitude caused by activation of A₁ adenosine receptors with N⁶-cyclopentyladenosine in the presence of the selective A_2A adenosine receptor agonist 2-p-(2-carboxyethy)phenethylamino-5′-N-ethylcarboxamidoadenosine (CGS 21680) in the rat hippocampus. In contrast, Dunwiddie et al. (1997) report that pre-treatment of hippocampal slices with CGS 21680 had no effect on subsequent responses to adenosine. Recently, Dixon et al. (1997) have shown the appearance of a low-affinity binding site for the adenosine A₁ receptor agonist 2-chloro-N-6-cyclopentyladenosine in the rat striatum caused by incubation of striatal synaptosomes with CGS 21680.

Although the aforementioned pre- and postsynaptic effects of adenosine have been well-characterised, little work has been carried out with regard to the effects of adenosine on the coupling between the two. This study investigates the effects of adenosine receptor agonists and antagonists on the population excitatory postsynaptic potential, the population spike, and the relationship between them i.e., EPSP–spike (E–S) coupling. The population excitatory postsynaptic potential (population EPSP) gives primarily a measure of membrane potential changes generated by excitatory synapses on the apical dendrites of CA1 pyramidal neurones. This extracellularly recorded potential has been shown to be affected by adenosine in the same way as intracellularly recorded EPSPs from the same cells (Proctor and Dunwiddie, 1987). The population spike reflects the summated firing of CA1 pyramidal neurones (Andersen et al., 1971) and gives a measure of the excitability of the postsynaptic neurone. E–S coupling gives an indication of the ability of a given level of synaptic depolarisation to induce the postsynaptic cell to fire an action potential. The three parameters measured can vary independently, and a description of all three is necessary to provide a full analysis of agents on neuronal function.