The Kölliker-Fuse nucleus orchestrates the timing of expiratory abdominal nerve bursting

Abstract

Coordination of respiratory pump and valve muscle activity is essential for normal breathing. A hallmark respiratory response to hypercapnia and hypoxia is the emergence of active exhalation, characterized by abdominal muscle pumping during the late one-third of expiration (late-E phase). Late-E abdominal activity during hypercapnia has been attributed to the activation of expiratory neurons located within the parafacial respiratory group (pFRG). However, the mechanisms that control emergence of active exhalation, and its silencing in restful breathing, are not completely understood. We hypothesized that inputs from the Kölliker-Fuse nucleus (KF) control the emergence of late-E activity during hypercapnia. Previously, we reported that reversible inhibition of the KF reduced postinspiratory (post-I) motor output to laryngeal adductor muscles and brought forward the onset of hypercapnia-induced late-E abdominal activity. Here we explored the contribution of the KF for late-E abdominal recruitment during hypercapnia by pharmacologically disinhibiting the KF in in situ decerebrate arterially perfused rat preparations. These data were combined with previous results and incorporated into a computational model of the respiratory central pattern generator. Disinhibition of the KF through local parenchymal microinjections of gabazine (GABA_A receptor antagonist) prolonged vagal post-I activity and inhibited late-E abdominal output during hypercapnia. In silico, we reproduced this behavior and predicted a mechanism in which the KF provides excitatory drive to post-I inhibitory neurons, which in turn inhibit late-E neurons of the pFRG. Although the exact mechanism proposed by the model requires testing, our data confirm that the KF modulates the formation of late-E abdominal activity during hypercapnia. NEW & NOTEWORTHY The pons is essential for the formation of the three-phase respiratory pattern, controlling the inspiratory-expiratory phase transition. We provide functional evidence of a novel role for the Kölliker-Fuse nucleus (KF) controlling the emergence of abdominal expiratory bursts during active expiration. A computational model of the respiratory central pattern generator predicts a possible mechanism by which the KF interacts indirectly with the parafacial respiratory group and exerts an inhibitory effect on the expiratory conditional oscillator.

Keywords: abdominal expiratory activity; active expiration; pons; respiratory pattern; ventral respiratory column.

Fig. 1.

Schematic of the brain stem respiratory network model including the inhibitory postinspiratory (post-I), excitatory postinspiratory [post-I(e)], and augmenting expiratory (aug-E) populations of the Bötzinger complex (BötC); the preinspiratory/inspiratory (pre-I/I) and early-inspiratory [early-I (1)] populations of the pre-Bötzinger complex (pre-BötC); the ramping inspiratory (ramp-I) and early-inspiratory [early-I (2)] populations of the rostral ventral respiratory group (rVRG); and the late-expiratory (late-E) population of the parafacial respiratory group (pFRG). This model includes excitatory drive elements, which are not modeled as populations and provide constant excitation to postsynaptic populations: drive from the pontine nuclei (Drive Pons), drive specifically from the Kölliker-Fuse nucleus (Drive KF), and drive from the retrotrapezoid nucleus (Drive RTN). Excitatory and inhibitory populations of 20–50 neurons are depicted as orange and blue elements, respectively. Similarly, projections from excitatory and inhibitory populations are colored orange and blue, respectively. Drive elements and projections associated with these elements are represented as green triangles or green arrows. The new additions to this model—Drive KF and its projections—are emphasized with black outline.

Fig. 2.

Functional and histological identification of the Kölliker-Fuse nucleus (KF). A: integrated recordings of abdominal (AbN), phrenic (PN), and cervical vagus (cVN) nerve activities from an in situ rat preparation, representative of the group, illustrating the respiratory responses to microinjections of glutamate (arrow) in the left (top) and right (bottom) sides of the KF. *represents an artifact generated during the removal of the injection micropipette. B: photomicrography of coronal section from the brain stem of a representative in situ rat preparation, illustrating the site of microinjection in the KF (arrow). C: schematic representations of all microinjection sites (black circles) into the KF (n = 6 each side). A7, A7 catecholaminergic cell group; DLL, dorsal nucleus of the lateral lemniscus; scp, superior cerebellar peduncle; s5, sensory root of trigeminal nerve; 4V, fourth ventricle.

Fig. 3.

Changes in baseline activities after microinjections of gabazine in the Kölliker-Fuse nucleus (KF). A: raw and integrated recordings of cervical vagus (cVN), phrenic (PN) and abdominal (AbN) nerve activities from a representative in situ rat preparation, illustrating the respiratory pattern before and after gabazine microinjections in the KF. B–E: average values of cVN post-I duration (normalized by expiratory time), coefficient of variation of PN burst frequency, mean PN burst frequency, and mean abdominal activity, respectively, before and after gabazine microinjections in the KF. *Different from baseline, P < 0.05. n = 6.

Fig. 4.

Disinhibition of the Kölliker-Fuse nucleus suppresses the generation of abdominal late-E activity during hypercapnia. Recordings, from representative in situ rat preparations, depict the activity of central vagus nerve (cVN), phrenic nerve (PN), and abdominal nerve (AbN) in eucapnia and hypercapnia, before treatment (A–E) and after gabazine microinjections (F–J). All recordings performed after gabazine microinjections are indicated in the light gray box. Filled arrows and open arrows in A and F indicate the beginning and end of the change in perfusate CO₂ fractional concentration from 5% to 8%, respectively. Zoomed-in traces B, D, G, and I depict respective preceding eucapnia epochs; C and H depict 8% hypercapnia; and E and J depict 10% hypercapnia. A–C and F–H are from a preparation challenged with 8% CO₂. D, E, I, and J come from a separate preparation that was challenged with 10% CO₂.

Fig. 5.

Disinhibition of the Kölliker-Fuse nucleus (KF) prevented the reduction in vagal postinspiratory activity and restrained the generation of late-E abdominal activity during hypercapnia. A and B: superimposed traces of integrated cervical vagus (cVN, gray) and abdominal nerve (AbN, black) activities during hypercapnia from a representative in situ rat preparation, before and after gabazine microinjections in the KF. Gray box indicates the inspiratory phase (coincident with phrenic burst), and arrows indicate the end of postinspiratory (post-I) activity in cVN. Note that before gabazine microinjections the onset of the late-E burst in AbN during hypercapnia is associated with a clear truncation in post-I activity. After gabazine microinjections, the amplitude of AbN late-E burst reduced and post-I cVN activity presented minor changes. C and D: average values of AbN late-E burst amplitude and variation of cVN post-I duration during hypercapnia, before and after gabazine microinjections in the KF. *Statistically significantly different from baseline, P < 0.05. n = 6.

Fig. 6.

Simulation of the glutamate microinjection into the Kölliker-Fuse nucleus (KF) by transient increase in drive from the KF to the respiratory CPG (arrow). Figure shows the activity of the phrenic nerve (PN) and the central vagus nerve (cVN) as well as the inhibitory postinspiratory (post-I), excitatory postinspiratory [post-I (e)], and augmenting-expiratory (aug-E) populations of the Bötzinger complex (BötC).

Fig. 7.

Simulation of the Kölliker-Fuse nucleus (KF) inhibition in eucapnia. Model activity of central pattern generator populations under eucapnia (A) and KF inhibition (B). Figure shows the activities of the preinspiratory/inspiratory (pre-I/I) population of the pre-Bötzinger complex (pre-BötC); the inhibitory postinspiratory (post-I), augmenting expiratory (aug-E), and excitatory postinspiratory [post-I (e)] populations of the Bötzinger complex (BötC); and the late-expiratory (late-E) population of the pFRG. Horizontal dashed line emphasizes the change in amplitude of the inhibitory post-I population between A and B.

Fig. 8.

Simulation of the Kölliker-Fuse nucleus (KF) inhibition and disinhibition in hypercapnia: motoneuron output in the model under hypercapnia + KF inhibition (A), hypercapnia (B), and hypercapnia + KF excitation (C), each depicting activity of the phrenic nerve (PN), the central vagus nerve (cVN), and the abdominal nerve (AbN).

Fig. 9.

Simulation of the Kölliker-Fuse nucleus (KF) inhibition and disinhibition in hypercapnia: model activity of central pattern generator populations under hypercapnia + KF inhibition (A), hypercapnia (B), and hypercapnia + KF excitation (C). post-I (BötC) traces are shaded gray for emphasis. Horizontal dashed black line in post-I (BötC) traces indicates the approximate threshold for activation of the late-E (pFRG). Vertical dashed gray lines indicate the phase of activation of late-E (pFRG). The included populations are the preinspiratory/inspiratory (pre-I/I) population of the pre-Bötzinger complex (pre-BötC); the inhibitory postinspiratory (post-I), augmenting expiratory (aug-E), and excitatory postinspiratory [post-I (e)] populations of the Bötzinger complex (BötC); and the late-expiratory (late-E) population of the pFRG.

Fig. 10.

Comparison of postinspiratory activity among 3 hypercapnia simulations. Activity of the post-I (e) (BötC) determines the postinspiratory component of the cVN burst. Vertical dashed line in each trace indicates the end of the postinspiratory burst.