Cholinergic suppression of heart rate acceleration during intrinsic optogenetic activation of sympathetic cardiac neurons in perfused hearts.

Cardiac autonomic pathways can be interrogated using optogenetics via selective expression of channelrhodopsin (ChR2) within intrinsic cardiac neurons. We tested the hypothesis that cholinergic activation via exogenous acetylcholine (ACh) would suppress the β-adrenergic heart rate (HR) acceleration induced by tyrosine hydroxylase (TH) neuron stimulation. HR responses were measured for wild-type (WT) mice at increasing norepinephrine (NE) concentrations and for transgenic mice that expressed ChR2 in TH neurons while illuminating the right atrium using a 465-nm microLED. Light pulse duty cycles (at 5 Hz) increasing from 5% to 50% caused proportional increases in HR; however, HR was unstable at duty cycles >30%. At 20% duty cycle, HR increased 39.43 ± 6.79%. HR increased for light pulse frequencies from 2.5 to 15 Hz (30 ms pulse width) but HR was unstable at 15 Hz. HR increases for photostimulated TH hearts and WT hearts perfused with NE (1-800 nM, WT + NE) had similar maxima but drastically different rise time constants (1.54 ± 0.08 vs. 18.04 ± 3.87 s, respectively). Increasing doses of ACh were added to the perfusate of WT + NE hearts and photostimulated TH hearts. High doses of ACh significantly diminished NE-induced and light-induced HR increases. Optogenetic activation of TH neurons caused rapid HR acceleration that was suppressed at higher ACh concentration than that required to suppress acceleration during NE perfusion. In WT + NE hearts, this occurred at 400 nM ACh and in TH hearts this did not occur until 1,800 nM ACh. Atrioventricular block occurred more rapidly in TH hearts during photostimulation as ACh concentration increased, demonstrating a time-dependent element in the interaction of simultaneous cholinergic and adrenergic activation.NEW & NOTEWORTHY Cardiac parasympathetic activity suppresses sympathetic activation, yet the details of the onset and magnitude of suppression are not fully understood. We found that higher doses of circulating ACh (cholinergic neurotransmitter) were required to suppress HR increases during optogenetic stimulation of adrenergic neurons in perfused mouse hearts compared with suppression after bolus administration of NE. We also found that optogenetic activation of adrenergic neurons after administering ACh caused severe AV block with an onset time that was dependent upon ACh concentration.