Article Figures & Data
Figures
- Fig. 1.
Schematic of method for measuring timing and amplitude of respiratory neuron modulation during the inspiratory burst. Top trace, Rectified ventral root activity (C2) averaged using ventral root burst onset as the trigger (vertical dashed line). Bottom trace, Type III inspiratory neuron activity triggered off ventral root burst. Duration (top horizontal arrow) and activity onset (second horizontal arrow) of inspiration-related activity in respiratory neurons are measured from C2 burst onset to half-maximal depolarization (types 1, 2, II, and III neurons) or hyperpolarization (pre-I neurons); these points fall on theline labeled half-height. Preinspiratory or postinspiratory deflections in membrane potential are measured relative to baseline membrane potential indicated by theline labeled resting Vm. Here, preinspiratory hyperpolarization onset and offset are measured relative to C2 burst onset (hyp duration, bottom, double arrow); the magnitude of hyperpolarization is measured relative to resting membrane potential (hyp amplitude,bottom, vertical arrow).
- Fig. 2.
A, Top, Rectified integrated C2 population activity from one experiment (τ = 20 msec). Bottom, Raster plot of periods from five experiments before (left of gray dashed line) and during (right of gray dashed line) phasic inflation. B, C, Pairedt tests on means reveal that phasic inflation significantly shortens burst duration (700 ± 80 vs 470 ± 50 msec; p < 0.01; B) and cycle period (9.5 ± 0.5 vs 7.5 ± 0.5 sec; p < 0.01;C).
- Fig. 3.
Biphasic neurons fire briskly during phasic inflation but are silent in control cycles. A, Raw traces of biphasic neuron membrane potential in control cycles (top) and with phasic inflation (bottom). In each panel, rectified integrated C2 ventral root activity (τ = 10 msec) is shown in the top trace, and biphasic neuron membrane potential is shown in the bottom trace. Transient inflations are indicated by gray boxes. The same format is used in Figures 4-8. B, Top, Average of biphasic neuron membrane potential over six cycles, triggered off ventral root inspiratory onset (vertical dashed line) with (black) and without (gray) phasic inflation in one biphasic neuron. Bottom, Bar graph of average inflation-induced firing duration in biphasic neurons (4 ± 0.8 sec; n = 7), scaled to thex-axis of the burst-triggered average. The same format is used in Figures 4-8. Note excitatory and inhibitory drive during inspiration in control cycles and brisk firing in the presence of phasic inflation. Both firing onset and firing offset of each neuron were calculated relative to ventral root inspiratory onset.C, Cycle-triggered histograms (30 bins per cycle) of two biphasic neurons. The first neuron fired tonically at low rates in control cycles (gray) and burst at a high rate during phasic inflation followed by slow adaptation afterward (dotted line), whereas the second neuron showed faster adaptation (solid line).
- Fig. 4.
Type 1 neurons are transiently inhibited during phasic inflation but show no change in preinspiratory activity.A, Raw traces of type 1 neuron membrane potential with and without phasic inflation are displayed as in Figure 3. In control cycles, type 1 neurons have a ramp-like depolarization during expiration and can begin to spike before ventral root inspiratory onset. Although preinspiratory trajectories with and without inflation coincide closely, in the presence of phasic inflation there is pronounced postinspiratory hyperpolarization coinciding with inflation (arrows). B, Burst-triggered average of seven cycles from one type 1 neuron, displayed as in Figure 3.Bar and column graphs show mean values for all eight type 1 neurons and are scaled to the axes of the averaged traces. Phasic inflation causes postinspiratory inhibition; theblack column to the right shows average postinspiratory inhibition in type 1 neurons (−5.8 ± 0.7 mV), which lasted 710 ± 100 msec (bottom, black bar). Phasic inflation significantly shortens (p < 0.01) type 1 inspiratory firing duration, from 1000 ± 120 msec (top, white bar) to 460 ± 60 msec (top, black bar).
- Fig. 5.
Preinspiratory depolarization in type 2 neurons begins earlier with phasic inflation, but postinspiratory activity is unchanged. A, Raw traces of type 2 neuron membrane potential with and without phasic inflation are displayed as in Figure 3. In control cycles, type 2 neuron membrane potential varies little during expiration and begins to depolarize before ventral root inspiratory onset. B, Burst-triggered average of seven cycles from one type 2 neuron, displayed as in Figure 3.Bar and column graphs show mean values for all three type 2 neurons and are scaled to the axes of the averaged traces. Although postinspiratory trajectories with and without inflation coincide closely, in the presence of phasic inflation, preinspiratory depolarization begins significantly earlier (p < 0.05; bottom,black bar) than in control cycles (bottom, white bar). Mean inflation-induced preinspiratory depolarization amplitude during phasic inflation (left, black column) was not significantly different from control (left, white column). Phasic inflation significantly shortens (p < 0.01) type 2 inspiratory firing duration, from 1160 ± 130 msec (top, white bar) to 670 ± 110 msec (top, black bar).
- Fig. 6.
Type II neuron inspiratory burst duration is shortened by phasic inflation, but preinspiratory and postinspiratory activity are unchanged. A, Raw traces of type II neuron membrane potential with and without phasic inflation are displayed as in Figure 3. In both control and phasic inflation cycles, type II neuron membrane potential is flat during expiration and rises abruptly during inspiration. B, Burst-triggered average of eight cycles from one type II neuron, displayed as in Figure 3.Bar graphs show mean values for all four type II neurons and are scaled to the axes of the averaged traces. In the averaged trace, the slope of inspiration-related depolarization remains nearly vertical, indicating tight coupling between motor output and type II activity. Firing onset during phasic inflation (bottom,black bar) and in control cycles (bottom,white bar) is unchanged. Preinspiratory membrane potential remained flat after application of bias currents to hold the cell at −87 mV; thus delayed onset to firing is not attributable to preinspiratory inhibition. Phasic inflation significantly shortens (p < 0.05) type II inspiratory firing duration, from 1010 ± 120 msec (top, white bar) to 640 ± 130 msec (top, black bar).
- Fig. 7.
Preinspiratory hyperpolarization begins earlier with phasic inflation in type III neurons. A, Raw traces of type III neuron membrane potential with and without phasic inflation are displayed as in Figure 3. In both control and phasic inflation cycles, type III neuron membrane potential shows consistent preinspiratory inhibition. B, Burst-triggered average of seven cycles from one type III neuron, displayed as in Figure 3. Bar and column graphs show mean values for all five type II neurons and are scaled to the axes of the averaged traces. Preinspiratory hyperpolarization onset occurs significantly earlier (p < 0.05) during phasic inflation (bottom, black bar) than in control cycles (bottom, white bar). Preinspiratory hyperpolarization amplitude (right,black column) and inspiratory burst onset are the same with and without phasic inflation. Phasic inflation significantly shortens (p < 0.01) type III inspiratory firing duration, from 1450 ± 170 msec (top,white bar) to 890 ± 80 msec (top,black bar).
- Fig. 8.
Preinspiratory depolarization begins earlier with phasic inflation in pre-I neurons, and postinspiratory firing is lost. A, Raw traces of pre-I neuron membrane potential with and without phasic inflation are displayed as in Figure3. In phasic inflation cycles, the characteristic preinspiratory and postinspiratory firing patterns of pre-I neurons in control cycles are transformed into pure preinspiratory firing. B, Burst-triggered average of nine cycles from one pre-I neuron, displayed as in Figure 3. Bar and column graphsshow mean values for all six pre-I neurons and are scaled to the axes of the averaged traces. Preinspiratory firing begins significantly earlier (p < 0.05) during phasic inflation (bottom, black bar) than in control cycles (bottom, white bar). Because no reversed outward currents are apparent after bias current application to hyperpolarize the cell to −75 mV, loss of postinspiratory activity is unlikely to be caused by Cl−-mediated inhibition.
- Fig. 9.
A, Mean onset times of maximal firing in type 1 (T-1), type 2 (T-2), type II (T-II), and type III (T-III) neurons or offset of firing in pre-I neurons. Onset times in control cycles (white columns) were not significantly different from onset times with phasic inflation (gray columns). Statistically significant differences in onset times between cell types were found, however, and are indicated by arrows between pairs of cell types. These differences support inferences about connectivity.B, Diagram of inferred connectivity between respiratory neurons. SAR afferent input (SAR) causes brisk firing of biphasic neurons (biphasic) after lung inflation. Postinspiratory inhibition seen only in type 1 neurons (Type 1) is consistent with biphasic neuron inhibition of type 1 neurons. Because type 1 inspiratory onset coincides closely with cervical ventral root motor output and fires earliest, type 1 neurons are hypothesized to drive inspiratory premotoneurons, such as type III (Type III) and motoneurons (MN). Delay in onset, together with low cycle-to-cycle variability in burst onset times in type II neurons (Type II), suggests an efference copy pathway from motoneurons to type II neurons. Because onset of type II neuron firing precedes but is not significantly different from onset of pre-I inhibition, type II neurons are hypothesized to inhibit pre-I neurons. Peri-inspiratory inhibition in type III neurons (Type III) is consistent with inhibition from pre-I neurons; the absence of this inhibition in type II neurons suggests that reciprocal inhibition from pre-I neurons to type II is not present. Because type III firing onset is significantly later than pre-I inhibition, reciprocal inhibition from type III neurons to pre-I neurons is not inferred. Neuron classes proposed to be causal to respiratory rhythm generation include pre-I neurons (Onimaru et al.; 1997), type 1 neurons (Rekling et al., 1996; Gray et al., 1999), and type 2 neurons (Thoby-Brisson et al.; 2000).
In this issue
