Calcium responses of circadian pacemaker neurons of the cockroach Rhyparobia maderae to acetylcholine and histamine

The accessory medulla (aMe) is the pacemaker that controls circadian activity rhythms in the cockroach Rhyparobia maderae. Not much is known about the classical neurotransmitters of input pathways to the cockroach circadian system. The circadian pacemaker center receives photic input from the compound eye, via unknown excitatory and GABAergic inhibitory entrainment pathways. In addition, neuropeptidergic inputs couple both pacemaker centers. A histamine-immunoreactive centrifugal neuron connects the ventral aMe with projection areas in the lateral protocerebrum and may provide non-photic inputs. To identify neurotransmitters of input pathways to the circadian clock with Fura-2-dependent Ca²⁺ imaging, primary cell cultures of the adult aMe were stimulated with acetylcholine (ACh), as the most prominent excitatory, and histamine, as common inhibitory neurotransmitter. In most of aMe neurons, ACh application caused dose-dependent increases in intracellular Ca²⁺ levels via ionotropic nicotinic ACh receptors. These ACh-dependent rises in Ca²⁺ were mediated by mibefradil-sensitive voltage-activated Ca²⁺ channels. In contrast, histamine application decreased intracellular Ca²⁺ levels in only a subpopulation of aMe cells via H2-type histamine receptor chloride channels. Thus, our data suggest that ACh is part of the light entrainment pathway while histamine is involved in a non-photic input pathway to the ventral circadian clock of the Madeira cockroach.