The most recently discovered adenosine receptor, the A3 receptor, belongs to the general class of G-protein coupled receptors, and has been linked to several putative effector mechanisms. One distinguishing characteristic of this receptor is that many adenosine agonists have affinities for the A3 receptor that are much lower than their corresponding affinities at the adenosine A1 receptor (Zhou et al., 1992), and this is true as well for the endogenous ligand, adenosine. It has been reported that A3 receptors are coupled in an inhibitory fashion to adenylyl cyclase (Zhou et al., 1992; Zhao et al., 1997), although in other systems, A3 receptor activation activates phospholipase C and leads to an elevation in inositol phosphate levels (Ali et al., 1990; Ramkumar et al., 1994; Auchampach et al., 1997), and this occurs in brain as well (Abbracchio et al., 1995). If this is the case, then increases in intracellular Ca2+ and activation of protein kinase C should occur as a consequence of agonist occupation of the A3 receptor. Recent studies in brain have suggested that A3 receptors can inhibit the function of presynaptic modulatory receptors (Dunwiddie et al., 1997; Macek et al., 1998), and that this effect is mediated via PKC (Macek et al., 1998; Diao and Dunwiddie, unpublished observations). However, other reports have proposed that postsynaptic A3 receptors in hippocampus may elicit responses that are linked to activation of PKA but not PKC (Fleming and Mogul, 1996), suggesting that a further consideration of effector pathways for this receptor is warranted. One response in hippocampal CA1 pyramidal neurons that is affected by increases in intracellular Ca2+, as well as activation of either PKC or PKA, is the K+-mediated afterhyperpolarization that follows activation of voltage dependent Ca2+ channels by a depolarizing stimulus. Previous studies have demonstrated that activation of β-adrenergic receptors will nearly abolish the AHP (Madison and Nicoll, 1982; Haas and Konnerth, 1983), an effect that can be mimicked directly by activation of PKA (Abdul-Ghani et al., 1996). Activation of PKC with phorbol esters will also inhibit this response (Malenka et al., 1986), and activation of either muscarinic receptors (Cole and Nicoll, 1984) or metabotropic glutamate receptors (Abdul-Ghani et al., 1996) reduce the AHP via increases in intracellular Ca2+ and subsequent activation of CaM-K. Furthermore, adenosine itself has been previously reported to either increase (Haas and Greene, 1984) or decrease (Dunwiddie, 1985) this current. Thus, this response appeared to be a likely candidate that would be sensitive to activation of multiple second messenger systems, but an effect of A3 receptor activation on this conductance has not been described.
The recent development of Cl-IB-MECA, an agonist that is approximately 2500-fold selective for the A3 vs. the A1 receptor, and 1400-fold selective for the A3 vs. the A2a receptor (Jacobson et al., 1995), has facilitated the study of the physiological consequences of selective activation of this receptor. Behavioral studies have demonstrated an A3 receptor-mediated depression of locomotor activity (Jacobson et al., 1993), but the effects of A3 receptor activation on neuronal activity at the cellular level are only poorly understood. A recent report has suggested that activation of A3 receptors can increase Ca2+ currents in hippocampal CA3 neurons (Fleming and Mogul, 1996), but apart from this, there have been few reports of specific actions of A3 receptor activation on neuronal activity.
Because the hippocampus and cerebellum show the highest levels of A3 mRNA in brain (De et al., 1993), and because the responses to A1 and A2 receptor activation have been well-characterized in this brain region (Dunwiddie, 1985; Greene and Haas, 1991; Cunha et al., 1994, 1995), we have investigated the electrophysiological actions of Cl-IB-MECA in this brain region.