Role of Synaptic Inhibition in the Coupling of the Respiratory Rhythms that Underlie Eupnea and Sigh Behaviors

Figures

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  Figure 1.. A, Field recording of preBötC activity, which includes inspiratory and sigh bursts. Average inspiratory cycle time (T¯) is calculated from six preceding cycle periods (Tϵ1...Tϵ6). Tϵ represents period of a single inspiratory cycle; TΔ represents the inspiratory-sigh interval; Tσ reflects the duration of the post-sigh apnea. B, A scatter plot showing z score transformation of burst area plotted against the post-sigh apnea duration, Tσ (normalized to the average inspiratory cycle time, T¯); z score distribution is calculated from all bursts from this slice.

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  Figure 2.. Variable timing of sigh breaths in vivo. Top, Histogram of the eupnea-sigh time intervals. Bin size is 0.016 s. Intervals near zero were pooled into one bin of size 0.048 s because distinct eupnea-related and sigh-related peaks in the signal could not be distinguished. Bottom, Sample sweeps from plethysmography recordings from one mouse. Lower case letters a, b, and c illustrate eupnea-sigh intervals that appear in corresponding bins plotted in the histogram above.

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  Figure 3.. Variable timing of sigh bursts in vitro. A, Histogram of normalized inspiratory-sigh intervals (TΔ/T¯) for all sigh bursts in all 13 rhythmically active slices under control conditions. The width of each bar is 0.1TΔ/T¯; μ marks the mean of the distribution, and κ denotes the 75th percentile line. Below are representative preBötC field recording traces depicting a variety of inspiratory-sigh intervals. Traces come from different slices. Lower case letters a, b, c, d, and e connect representative traces with their position on the inspiratory-sigh interval histogram above. B, C, Identical inspiratory-sigh interval analysis in the presence of 5 μm strychnine (B) as well as both 5 μm strychnine and 5 μm picrotoxin (C).

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  Figure 4.. Cumulative probability histograms showing the normalized inspiratory-sigh interval (TΔ/T¯) under control conditions (blue), 5 μm strychnine (orange), as well as 5 μm strychnine and 5 μm picrotoxin (red); κ marks the 75th percentile line for each distribution, distinguished by color. This analysis pools all sighs measured across all 13 slices.

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  Figure 5.. Average relative duration of the post-sigh apnea in control (blue), 5 μm strychnine (orange), as well as 5 μm strychnine and 5 μm picrotoxin (red). Each dot represents the average normalized post-sigh apnea for all sighs recorded in one slice. Horizontal bars represent the average of all slices in each condition.

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  Figure 6.. Chloride-mediated synapses are inhibitory in neonatal preBötC neurons. A, B, Image of Dbx1;Ai148 slice preparation under Dodt contrast (A) and fluorescence microscopy (B). preBötC (red box) colocalized with the NAsc and the principle loop of the inferior olive (IOloop). Scale bar for both A, B: 500 μm. C, Voltage trajectory of a Dbx1 preBötC neuron during formation of a gramicidin perforated patch, firing in sync with XII nerve activity. Dodt image shows the patch pipette on the neuronal soma. The magenta arrow (labeled p.) points to the patch pipette, the cyan arrow (labeled soma) indicates the patched cell body. GCaMP6f image shows the neuron was Dbx1 derived. The lack of fluorescence in the Alexa Fluor 568 image verifies the integrity of the perforated patch. D, Membrane voltage trajectory of the same neuron during blockade of chloride-mediated synaptic inhibition via addition of 5 μm strychnine and 5 μm picrotoxin in the bath. Alexa Fluor 568 continues to verify the integrity of the perforated patch. E, The same neuron in a whole-cell patch configuration shows Alexa Fluor 568 enters the soma from the patch pipette. The Dbx1 neuron is still rhythmically active and fires in…

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  Figure 7.. Meta-analysis of ECl during development in hindbrain and spinal cord of mice and rats. Red reflects rat data, black reflects mouse. The abscissa shows age (in days) centered at birth (P0). The ordinate shows ECl (in mV) as a function of age. Below the x-axis, we show binary classification of ECl as either above (excitatory) or below (inhibitory) baseline membrane potential as a function of developmental age. Our present data are labeled Borrus et al. (2020). We obtained ECl data from perinatal mice from: Branchereau et al. (2000), Delpy et al., 2008, Ritter and Zhang (2000), and data from the present manuscript, i.e., Borrus et al. (2020). We obtained ECl data from perinatal rats from: Brockhaus and Ballanyi (1998), Fregosi et al. (2004), Ren and Greer, 2006, as well as Shao and Feldman (1997).