Structure-activity relationships of BnOCPA derivatives at native pre- and postsynaptic adenosine A₁Rs

The addition of a benzyloxy group to the adenosine A₁R agonist cyclopentyladenosine (CPA) produces a compound (BnOCPA) that selectivity activates A₁R signalling via the G protein alpha subunit Gob. In rat hippocampus, BnOCPA inhibits CA3-CA1 synaptic transmission (IC₅₀ 65 nM) likely via Gob, with no effect on Goa-mediated postsynaptic membrane electrophysiology at concentrations up to 1 μM. We have investigated how modifications of the benzyloxy group of BnOCPA affects signalling: addition of methoxy group, MBnOCPA, substituting a phenoxy group, PhOCPA; and addition of bromide to the phenoxy group, BPhOCPA. These derivatives, previously shown to be selective A₁R agonists, were assessed through inhibition of hippocampal CA3-CA1 synaptic transmission and postsynaptic effects on CA1 pyramidal neurons. All compounds inhibited synaptic transmission and increased paired-pulse facilitation, indicative of presynaptic actions. At a concentration that inhibited synaptic transmission (>75 %), none of the compounds hyperpolarised the resting membrane potential and thus retained the pre- vs postsynaptic selectivity of BnOCPA. Increasing their concentration allowed a measure of the selectivity range. BnOCPA hyperpolarised ∼ 40 % of neurons at 3 μM (∼ 50x the IC₅₀ for synaptic inhibition). For PhOCPA and MBnOCPA, hyperpolarisation, occurred for some neurons, at 1 μM. For BPhOCPA, no hyperpolarisation occurred at concentrations up to 10 μM. Pre-application of the derivatives (at 300 nM) reduced adenosine-mediated hyperpolarisation, showing they bind but do not activate postsynaptic A₁Rs. This data shows that modifying the structure of BnOCPA changes the discrimination between different native A₁Rs, likely based upon selective activation of G-protein coupling.