Noradrenergic tuning, not simple rate effects, produces temperature‐sensitivity of the respiratory network in bullfrogs

Abstract

Animals match ventilation to metabolic and acid‐base regulatory demands during changes in body temperature. Across vertebrates, ventilation increases at warmer temperatures and decreases at cooler temperatures. At the organismal level this is attributed to metabolic feedback and/or alpha‐stat pH regulation; however, the cellular mechanisms that produce ventilatory output in response to brain temperature changes remain unclear. To identity mechanisms underlying temperature‐sensitivity of the respiratory control network, we used brainstem‐spinal cord preparations producing spontaneously active, rhythmic motor output similar to breathing in vivo of adult bullfrogs Lithobates catesbeianus. In vitro brainstem‐spinal cords were superfused with artificial cerebrospinal fluid (aCSF) equilibrated at 90% O2, 1.3% CO2, and balance N2. Whole nerve recordings from the trigeminal (V) and vagus (X) nerves were used for measuring respiratory‐related activity. We applied temperature ramps from 20°C to 15°C and then to 25°C; each step lasted 15 minutes. Bursting frequency was analyzed for the last 5 minutes of each step and then normalized to percent of baseline (20°C). Consistent with in vivo and in vitro data (Bícego‐Nahas and Branco, 1999; Morales and Hedrick, 2002), we demonstrate that the frequency of respiratory‐related nerve activity is stable across high temperatures, but not lower temperatures (One‐way ANOVA p=0.0013; percent of baseline significantly lower at 15°C compared to 20°C and compared to 25°C, but no difference between 20°C and 25°C; Tukey's Multiple Comparison Test). The locus coeruleus (LC) is a nucleus of the respiratory network and is the main supplier of norepinephrine in the brain. LC neurons from bullfrogs are paradoxically activated by decreases in temperature (Santin et al., 2013) suggesting that firing frequencies inversely proportional to temperature may play a role in setting the respiratory frequency across temperatures. To identify the role of norepinephrine in generating the respiratory frequency, we applied the temperature protocol while blocking the main adrenergic receptors (AR). For blocking α1AR, preparations were superfused with aCSF containing Prazosin and for blocking α2AR we used RX821002. We found that bursting stability at high temperatures is disrupted when α1AR are blocked (One‐way ANOVA p=0.0011; percent of baseline significantly lower at 15°C compared to 20°C and 25°C, and percent of baseline significantly lower at 20°C compared to 25°C; Tukey's Multiple Comparison Test). In contrast, the inhibition of bursting frequency at 15°C was lost when α2AR were blocked (One‐way ANOVA p=0.1649; percent of baseline not different at 15°C, 20°C and 25°C). These results imply that norepinephrine tuning through different receptors, rather than simple Q10 effects, plays a major role in generating the breathing frequency across temperatures to match metabolic demands and acid‐base regulation requirements at those temperatures.Support or Funding InformationSupport for this project was provided by the Department of Biological Sciences at Wright State University as well as a Biology Award for Research Excellence and a Graduate Student Assembly Original Research Grant (both to M.V.).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.