Single cell laser dissection with molecular beacon polymerase chain reaction identifies 2A as the predominant serotonin receptor subtype in hypoglossal motoneurons

We hypothesize that sleep state-dependent withdrawal of serotonin (5-hydroxytryptamine, 5-HT) at upper airway (UAW) dilator motoneurons contributes significantly to sleep-related suppression of dilator muscle activity in obstructive sleep apnea. Identification of 5-HT receptor subtypes involved in postsynaptic facilitation of UAW motoneuron activity may provide pharmacotherapies for this prevalent disorder. We have adapted two assays to provide semi-quantitative measurements of mRNA copy numbers for 5-HT receptor subtypes in single UAW motoneurons. Specifically, soma of 111 hypoglossal (XII) motoneurons in 10 adult male rats were captured using a laser dissection microscope, and then used individually in single round molecular beacon polymerase chain reaction (PCR) for real-time quantitation of 5-HT 2A, 5-HT 2C, 5-HT 3, 5-HT 4, 5-HT 5A, 5-HT 5B, 5-HT 6 or 5-HT 7 receptor. Receptor mRNA copy numbers from single XII motoneurons were compared to control samples from within the XII nucleus and lateral medulla. All 20 motoneuronal soma assayed for the 5-HT 2A receptor had measurable copy numbers (7028±2656 copies/cell). In contrast, copy numbers for the 5-HT 2A receptor in XII non-motoneuronal ( n=17) and lateral medulla ( n=15) samples were 81±51 copies and 83±35 copies, respectively, P