The effects of 5′- N-ethylcarboxamidoadenosine on evoked release of [ 3H]serotonin in the rat nucleus tractus solitarius

Abstract

5′- N-Ethylcarboxamidoadenosine (NECA) a non-selective adenosine A 1 and A 2a receptor agonist was employed in stimulated (3 Hz, 25 mA, 1 min) superfused nucleus tractus solitarius (NTS) brain slices loaded with [ 3H]5-hydroxytryptamine ([ 3H]5-HT), and ligand binding with [ 3H]NECA on NTS membranes. Superfusion of NTS slices with 1.0 and 3.0 nM NECA for 5 min prior to S 2 stimulation produced an augmented release of [ 3H]5-HT (35.7%) above the control S 2 S 1 ratio. When the duration of NECA perfusion was increased to 20 min prior to S 2, the S 2 S 1 ratio was depressed 21% at 1.0 nM for [ 3H]5-HT release. The augmented release of [ 3H]5-HT by NECA at 5 min was blocked by the adenosine A 2a antagonist 8-(3-chlorostyryl)caffeine (CSC; 100 nM), and was reduced but not blocked by the A 1 antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; 10 nM). Saturation binding assays with [ 3H]NECA on NTS membranes showed two binding sites, a high affinity site with a K D 2.18 nM and low affinity site with a K D of 44.9 nM. With the selective adenosine A 2a antagonist CSC the high affinity site was blocked while 10 nM DPCPX, the A 1 antagonist, reduced binding of the low affinity site, but did not abolish it. NECA binds to two different adenosine receptor sites in NTS membranes with the A 2a receptor being the high affinity site. The same A 2a site is associated with the augmented neurotransmitter release of [ 3H]5-HT with 5 min tissue exposure since it is blocked by CSC. Longer tissue exposure to NECA resulted in desensitization and finally inhibition of release possibly associated with adenosine A 1 receptors.