Nucleus Retroambiguus Research Articles

Central Autonomic Mechanisms Involved in the Control of Laryngeal Activity and Vocalization.

In humans, speech is a complex process that requires the coordinated involvement of various components of the phonatory system, which are monitored by the central nervous system. The larynx in particular plays a crucial role, as it enables the vocal folds to meet and converts the exhaled air from our lungs into audible sounds. Voice production requires precise and sustained exhalation, which generates an air pressure/flow that creates the pressure in the glottis required for voice production. Voluntary vocal production begins in the laryngeal motor cortex (LMC), a structure found in all mammals, although the specific location in the cortex varies in humans. The LMC interfaces with various structures of the central autonomic network associated with cardiorespiratory regulation to allow the perfect coordination between breathing and vocalization. The main subcortical structure involved in this relationship is the mesencephalic periaqueductal grey matter (PAG). The PAG is the perfect link to the autonomic pontomedullary structures such as the parabrachial complex (PBc), the Kölliker-Fuse nucleus (KF), the nucleus tractus solitarius (NTS), and the nucleus retroambiguus (nRA), which modulate cardiovascular autonomic function activity in the vasomotor centers and respiratory activity at the level of the generators of the laryngeal-respiratory motor patterns that are essential for vocalization. These cores of autonomic structures are not only involved in the generation and modulation of cardiorespiratory responses to various stressors but also help to shape the cardiorespiratory motor patterns that are important for vocal production. Clinical studies show increased activity in the central circuits responsible for vocalization in certain speech disorders, such as spasmodic dysphonia because of laryngeal dystonia.

A brainstem circuit for phonation and volume control in mice

Mammalian vocalizations are critical for communication and are produced through the process of phonation, in which expiratory muscles force air through the tensed vocal folds of the larynx, which vibrate to produce sound. Despite the importance of phonation, the motor circuits in the brain that control it remain poorly understood. In this study, we identified a subpopulation of ~160 neuropeptide precursor Nts (neurotensin)-expressing neurons in the mouse brainstem nucleus retroambiguus (RAm) that are robustly activated during both neonatal isolation cries and adult social vocalizations. The activity of these neurons is necessary and sufficient for vocalization and bidirectionally controls sound volume. RAm Nts neurons project to all brainstem and spinal cord motor centers involved in phonation and activate laryngeal and expiratory muscles essential for phonation and volume control. Thus, RAm Nts neurons form the core of a brain circuit for making sound and controlling its volume, which are two foundations of vocal communication.

Large-Scale Mapping of Vocalization-Related Activity in the Functionally Diverse Nuclei in Rat Posterior Brainstem.

The identity and location of vocalization pattern generating (VPG) circuits in mammals is debated. Based on physiological experiments, investigators suggested anterior brainstem circuits in the reticular formation, and anatomic evidence suggested the nucleus retroambiguus (NRA) in the posterior brainstem, or combinations of these sites as the putative mammalian VPG. Additionally, vocalization loudness is a critical factor in acoustic communication. However, many of the underlying neuronal mechanisms are still unknown. Here, we evoked calls by stimulation of the periaqueductal gray in anesthetized male rats, performed a large-scale mapping of vocalization-related activity using the activity marker c-fos, and high-density recordings of brainstem circuits using Neuropixels probes. Both c-fos expression and recording of vocalization-related activity point to a participation of the NRA in vocalization. More important, among our recorded structures, we found that the NRA is the only brainstem area showing a strong correlation between unit activity and call intensity. In addition, we observed functionally diverse patterns of vocalization-related activity in a set of regions around NRA. Dorsal to NRA, we observed activity specific to the beginning and end of vocalizations in the posterior level of the medullary reticular nucleus, dorsal part, whereas medial and lateral to the NRA, we observed activity related to call initiation. No clear vocalization-related activity was observed at anterior brainstem sites. Our findings suggest a set of functionally heterogeneous regions around the NRA contribute to vocal pattern generation in rats.SIGNIFICANCE STATEMENT Vocalization patterns are shaped in the mammalian brainstem, but the identity and location of the circuits involved is debated. Additionally, the neuronal mechanisms of vocal intensity control are still unknown. This study consisted of a large-scale mapping of brainstem vocalization circuits based on the activity marker c-fos and high-density recordings with Neuropixels probes. The results confirm the role of nucleus retroambiguus in call production and point to a key role of neurons in this nucleus in loudness control. Dorsal to the nucleus retroambiguus and in the posterior medulla, the authors identify neurons with activity specific to the beginning and end of vocalizations. The results point to specific neural dials for various aspects of rat vocalization control in the posterior brainstem.

Reciprocal connectivity of the periaqueductal gray with the ponto-medullary respiratory network in rat

Synaptic activities of the periaqueductal gray (PAG) can modulate or appropriate the respiratory motor activities in the context of behavior and emotion via descending projections to nucleus retroambiguus. However, alternative anatomical pathways for the mediation of PAG-evoked respiratory modulation via core nuclei of the brainstem respiratory network remains only partially described. We injected the retrograde tracer Cholera toxin subunit B (CT-B) in the pontine Kölliker-Fuse nucleus (KFn, n = 5), medullary Bötzinger (BötC, n = 3) and pre-Bötzinger complexes (pre-BötC; n = 3), and the caudal raphé nuclei (n = 3), and quantified the descending connectivity of the PAG targeting these brainstem respiratory regions. CT-B injections in the KFn, pre-BötC, and caudal raphé, but not in the BötC, resulted in CT-B-labeled neurons that were predominantly located in the lateral and ventrolateral PAG columns. In turn, CT-B injections in the lateral and ventrolateral PAG columns (n = 4) produced the highest numbers of CT-B-labeled neurons in the KFn and far fewer numbers of labeled neurons in the pre-BötC, BötC, and caudal raphé. Analysis of the relative projection strength revealed that the KFn shares the densest reciprocal connectivity with the PAG (ventrolateral and lateral columns, in particular). Overall, our data imply that the PAG may engage a distributed respiratory rhythm and pattern generating network beyond the nucleus retroambiguus to mediate downstream modulation of breathing. However, the reciprocal connectivity of the KFn and PAG suggests specific roles for synaptic interaction between these two nuclei that are most likely related to the regulation of upper airway patency during vocalization or other volitional orofacial behaviors.

A Functionally and Anatomically Bipartite Vocal Pattern Generator in the Rat Brain Stem.

SummaryThe mammalian vocal pattern generator is situated in the brainstem but its exact structure is debated. We mapped these circuits in rats by cooling and microstimulation. Local cooling disrupted call production above an anterior and a posterior brainstem position. Anterior cooling affected predominantly high-frequency calls, whereas posterior cooling affected low-frequency calls. Electrical microstimulation of the anterior part led to modulated high-frequency calls, whereas microstimulation of the posterior part led to flat, low-frequency calls. At intermediate positions cooling did not affect calls and stimulation did not elicit calls. The anterior region corresponds to a subsection of the parvicellular reticular formation that we term the vocalization parvicellular reticular formation (VoPaRt). The posterior vocalization sites coincide with the nucleus retroambiguus (NRA). VoPaRt and NRA neurons were very small and the VoPaRt was highly myelinated, suggestive of high-speed processing. Our data suggest an anatomically and functionally bipartite vocal pattern generator.

Microstimulation in Different Parts of the Periaqueductal Gray Generates Different Types of Vocalizations in the Cat

In the cat four different types of vocalization, mews, howls, cries, and hisses were generated by microstimulation in different parts of the periaqueductal gray (PAG). While mews imply positive vocal expressions, howls, hisses, and cries represent negative vocal expressions. In the intermediate PAG, mews were generated in the lateral column, howls, and hisses in the ventrolateral column. Cries were generated in two other regions, the lateral column of the rostral PAG and the ventrolateral column of the caudal PAG. In order to define the specific motor patterns of the mews, howls, and cries, the following muscles were recorded during these vocalizations; larynx (cricothyroid, thyroarytenoid, and posterior cricoarytenoid), tongue (genioglossus), jaw (digastric), and respiration muscles (diaphragm, internal intercostal, external, and internal abdominal oblique). During these mews, howls, and cries we analyzed the frequency, intensity, activation cascades power density, turns, and amplitude analysis of the electromyograms (EMGs). It appeared that each type of vocalization consists of a specific circumscribed motor coordination. The nucleus retroambiguus (NRA) in the caudal medulla is known to serve as the final premotor interneuronal output system for vocalization. Although neurochemical microstimulation in the NRA itself also generated vocalizations, they only consisted of guttural sounds, the EMGs of which involved only small parts of the EMGs of the mews, howls, and cries generated by neurochemical stimulation in the PAG. These results demonstrate that positive and negative vocalizations are generated in different parts of the PAG. These parts have access to different groups of premotoneurons in the NRA, that, in turn, have access to different groups of motoneurons in the brainstem and spinal cord, resulting in different vocalizations. The findings would serve a valuable model for diagnostic assessment of voice disorders in humans.

Effects of pharmacological lesion of the nucleus retroambiguus region on the pharyngeal phase of swallowing

Pharyngeal swallowing is controlled by synaptic interactions within a swallowing central pattern generator (sw-CPG) that is composed of a dorsal and a ventral swallowing group (VSG). Here, we used electrical stimulation (10 s) of the superior laryngeal nerve (SLN; 20 Hz; pulse width: 100 μs) to explore the role of the VSG in an arterially-perfused brainstem preparation of rats. To investigate the effects of pharmacological lesion (local microinjection of an GABA(A)-R agonist) of the nucleus retroambiguus (NRA), a designated component of the VSG, we recorded phrenic (PNA) and vagal nerve (VNA) activities. Control SLN stimulation with stepwise increasing stimulus intensities (from 20 μA to 160 μA) elicited robust suppression of PNA and evoked sequential swallowing activity in the VNA. Lesioning of the NRA had no effect on the pattern of pharyngeal swallowing, but significantly increased the sensory gating of SLN inputs. We conclude that the NRA is not part of the VSG, but appears to have important roles for the central gating of swallowing.

Involvement of rat posterior prelimbic and cingulate area 2 in vocalization control.

Microstimulation mapping identified vocalization areas in primate anterior cingulate cortex. Rat anterior cingulate and medial prefrontal areas have also been intensely investigated, but we do not know, how these cortical areas contribute to vocalizations and no systematic mapping of stimulation‐evoked vocalizations has been performed. To address this question, we mapped microstimulation‐evoked (ultrasonic) vocalizations in rat cingulate and medial prefrontal cortex. The incidence of evoked vocalizations differed markedly between frontal cortical areas. Vocalizations were most often evoked in posterior prelimbic cortex and cingulate area 2, whereas vocalizations were rarely evoked in dorsal areas (vibrissa motor cortex, secondary motor cortex and cingulate area 1) and anterior areas (anterior prelimbic, medial‐/ventral‐orbital cortex). Vocalizations were observed at intermediate frequencies in ventro‐medial areas (infralimbic and dorsopeduncular cortex). Various complete, naturally occurring calls could be elicited. In prelimbic cortex superficial layer microstimulation evoked mainly fear calls with low efficacy, whereas deep layer microstimulation evoked mainly 50 kHz calls with high efficacy. Vocalization stimulation thresholds were substantial (70–500 μA, the maximum tested; on average ~400 μA) and latencies were long (median 175 ms). Posterior prelimbic cortex projected to numerous targets and innervated brainstem vocalization centers such as the intermediate reticular formation and the nucleus retroambiguus disynaptically via the periaqueductal gray. Anatomical position, stimulation effects and projection targets of posterior prelimbic cortex were similar to that of monkey anterior cingulate vocalization cortex. Our data suggest that posterior prelimbic cortex is more closely involved in control of vocalization initiation than in specifying acoustic details of vocalizations.

A Specialized Neural Circuit Gates Social Vocalizations in the Mouse

Vocalizations are fundamental to mammalian communication, but the underlying neural circuits await detailed characterization. Here, we used an intersectional genetic method to label and manipulate neurons in the midbrain periaqueductal gray (PAG) that are transiently active in male mice when they produce ultrasonic courtship vocalizations (USVs). Genetic silencing of PAG-USV neurons rendered males unable to produce USVs and impaired their ability to attract females. Conversely, activating PAG-USV neurons selectively triggered USV production, even in the absence of any female cues. Optogenetic stimulation combined with axonal tracing indicates that PAG-USV neurons gate downstream vocal-patterning circuits. Indeed, activating PAG neurons that innervate the nucleus retroambiguus, but not those innervating the parabrachial nucleus, elicited USVs in both male and female mice. These experiments establish that a dedicated population of PAG neurons gives rise to a descending circuit necessary and sufficient for USV production while also demonstrating the communicative salience of male USVs. VIDEO ABSTRACT.

Tauopathy in the periaqueductal gray, kölliker-fuse nucleus and nucleus retroambiguus is not predicted by ultrasonic vocalization in tau-P301L mice

Upper airway and vocalization control areas such as the periaqueductal gray (PAG), kölliker-fuse nucleus (KF) and nucleus retroambiguus (NRA) are prone to developing tauopathy in mice expressing the mutant human tau P301L protein. Consequently, impaired ultrasonic vocalization (USV) previously identified in tau-P301L mice at the terminal disease stage of 8–9 months of age, was attributed to the presence of tauopathy in these regions. Our aim was to establish whether the onset of USV disorders manifest prior to the terminal stage, and if USV disorders are predictive of the presence of tauopathy in the PAG, KF and NRA. USVs produced by tau-P301L and wildtype mice aged 3–4, 5–6 or 8–9 months were recorded during male-female interaction. Immunohistochemistry was then performed to assess the presence or degree of tauopathy in the PAG, KF and NRA of mice displaying normal or abnormal USV patterns. Comparing various USV measurements, including the number, duration and frequency of calls, revealed no differences between tau-P301L and wildtype mice across all age groups, and linear discriminant analysis also failed to identify separate USV populations. Finally, the presence of tauopathy in the PAG, KF and NRA in individual tau-P301L mice did not reliably associate with USV disorders. Our findings that tauopathy in designated mammalian vocalization centres, such as the PAG, KF and NRA, did not associate with USV disturbances in tau-P301L mice questions whether USV phenotypes in this transgenic mouse are valid for studying tauopathy-related human voice and speech disorders.

Expression of the transcription factor FOXP2 in brainstem respiratory circuits of adult rat is restricted to upper-airway pre-motor areas

Expression of the transcription factor FOXP2 is linked to brain circuits that control motor function and speech. Investigation of FOXP2 protein expression in respiratory areas of the ponto-medullary brainstem of adult rat revealed distinct rostro-caudal expression gradients. A high density of FOXP2 immunoreactive nuclei was observed within the rostral pontine Kölliker-Fuse nucleus, compared to low densities in caudal pontine and rostral medullary respiratory nuclei, including the: (i) noradrenergic A5 and parafacial respiratory groups; (ii) Bötzinger and pre-Bötzinger complex and; (iii) rostral ventral respiratory group. Moderate densities of FOXP2 immunoreactive nuclei were observed in the caudal ventral respiratory group and the nucleus retroambiguus, with significant density levels found in the caudal half of the dorsal respiratory group and the hypoglossal pre-motor area lateral around calamus scriptorius. FOXP2 immunoreactivity was absent in all cranial nerve motor nuclei. We conclude that FOXP2 expression in respiratory brainstem areas selectively delineates laryngeal and hypoglossal pre-motor neuron populations essential for the generation of sound and voice.

The physiological motor patterns produced by neurons in the nucleus retroambiguus in the rat and their modulation by vagal, peripheral chemosensory, and nociceptive stimulation.

The nucleus retroambiguus (NRA) is a neuronal cell group in the medullary ventrolateral tegmentum, rostrocaudally between the obex and the first cervical spinal segment. NRA neurons are premotor interneurons with direct projections to the motoneurons of soft palate, pharynx, and larynx in the nucleus ambiguus in the lateral medulla as well as to the motoneurons in the spinal cord innervating diaphragm, abdominal, and pelvic floor muscles and the lumbosacral motoneurons generating sexual posture. These NRA premotor interneurons receive very strong projections from the periaqueductal gray (PAG) in the context of basic survival mechanisms as fight, flight, freezing, sound production, and sexual behavior. In the present study in rat we investigated the physiological motor patterns generated by NRA neurons, as the result of vagal, peripheral chemosensory, and nociceptive stimulation. The results show that the NRA contains phasic respiratory modulated neurons, as well as nonphasic tonically modulated neurons. Stimulation in the various rostrocaudal levels of the NRA generates site-specific laryngeal, respiratory, abdominal, and pelvic floor motor activities. Vagal and peripheral chemosensory stimulation induces both excitatory and inhibitory modulation of phasic NRA-neurons, while peripheral chemosensory and nociceptive stimulation causes excitation and inhibition of nonphasic NRA-neurons. These results are in agreement with the concept that the NRA represents a multifunctional group of neurons involved in the output of the emotional motor system, such as vomiting, vocalization, mating, and changes in respiration.

Two different motor systems are needed to generate human speech

Vocalizations such as mews and cries in cats or crying and laughter in humans are examples of expression of emotions. These vocalizations are generated by the emotional motor system, in which the mesencephalic periaqueductal gray (PAG) plays a central role, as demonstrated by the fact that lesions in the PAG lead to complete mutism in cats, monkeys, as well as in humans. The PAG receives strong projections from higher limbic regions and from the anterior cingulate, insula, and orbitofrontal cortical areas. In turn, the PAG has strong access to the caudal medullary nucleus retroambiguus (NRA). The NRA is the only cell group that has direct access to the motoneurons involved in vocalization, i.e., the motoneuronal cell groups innervating soft palate, pharynx, and larynx as well as diaphragm, intercostal, abdominal, and pelvic floor muscles. Together they determine the intraabdominal, intrathoracic, and subglottic pressure, control of which is necessary for generating vocalization. Only humans can speak, because, via the lateral component of the volitional or somatic motor system, they are able to modulate vocalization into words and sentences. For this modulation they use their motor cortex, which, via its corticobulbar fibers, has direct access to the motoneurons innervating the muscles of face, mouth, tongue, larynx, and pharynx. In conclusion, humans generate speech by activating two motor systems. They generate vocalization by activating the prefrontal‐PAG‐NRA‐motoneuronal pathway, and, at the same time, they modulate this vocalization into words and sentences by activating the corticobulbar fibers to the face, mouth, tongue, larynx, and pharynx motoneurons. J. Comp. Neurol. 524:1558–1577, 2016. © 2015 Wiley Periodicals, Inc.

Special edition on positive emotions.

Special edition on positive emotions.

Two different motor systems are needed to generate human speech

Vocalizations such as mews and cries in cats or crying and laughter in humans are examples of expression of emotions. These vocalizations are generated by the emotional motor system, in which the mesencephalic periaqueductal gray (PAG) plays a central role, as demonstrated by the fact that lesions in the PAG lead to complete mutism in cats, monkeys, as well as in humans. The PAG receives strong projections from higher limbic regions and from the anterior cingulate, insula, and orbitofrontal cortical areas. In turn, the PAG has strong access to the caudal medullary nucleus retroambiguus (NRA). The NRA is the only cell group that has direct access to the motoneurons involved in vocalization, i.e., the motoneuronal cell groups innervating soft palate, pharynx, and larynx as well as diaphragm, intercostal, abdominal, and pelvic floor muscles. Together they determine the intraabdominal, intrathoracic, and subglottic pressure, control of which is necessary for generating vocalization. Only humans can speak, because, via the lateral component of the volitional or somatic motor system, they are able to modulate vocalization into words and sentences. For this modulation they use their motor cortex, which, via its corticobulbar fibers, has direct access to the motoneurons innervating the muscles of face, mouth, tongue, larynx, and pharynx. In conclusion, humans generate speech by activating two motor systems. They generate vocalization by activating the prefrontal‐PAG‐NRA‐motoneuronal pathway, and, at the same time, they modulate this vocalization into words and sentences by activating the corticobulbar fibers to the face, mouth, tongue, larynx, and pharynx motoneurons. J. Comp. Neurol. 524:1558–1577, 2016. © 2015 Wiley Periodicals, Inc.

Motor organization of positive and negative emotional vocalization in the cat midbrain periaqueductal gray.

Neurochemical microstimulation in different parts of the midbrain periaqueductal gray (PAG) in the cat generates four different types of vocalization, mews, howls, cries, and hisses. Mews signify positive vocal expression, whereas howls, hisses, and cries signify negative vocal communications. Mews were generated in the lateral column of the intermediate PAG and howls and hisses in the ventrolateral column of the intermediate PAG. Cries were generated in two regions, the lateral column of the rostral PAG and the ventrolateral column of the caudal PAG. To define the specific motor patterns belonging to mews, howls, and cries, the following muscles were recorded during these vocalizations: larynx (cricothyroid, thyroarytenoid, and posterior cricoarytenoid), tongue (genioglossus), jaw (digastric), and respiration (diaphragm, internal intercostal, external abdominal oblique, and internal abdominal oblique) muscles. Furthermore, the frequency, intensity, activation cascades, and turns and amplitude analyses of the electromyograms (EMGs) during these vocalizations were analyzed. The results show that each type of vocalization consists of a specific, circumscribed motor coordination. The nucleus retroambiguus (NRA) in the caudal medulla serves as the final premotor interneuronal output system for vocalization. NRA neurochemical microstimulation also generated vocalizations (guttural sounds). Analysis of the EMGs demonstrated that these vocalizations consist of only small parts of the emotional voalizations generated by neurochemical stimulation in the PAG. These results demonstrate that motor organization of positive and negative emotional vocal expressions are segregated in the PAG and that the PAG uses the NRA as a tool to gain access to the motoneurons generating vocalization.

Novel Insights into Anatomical and Functional Roles of Nucleus Retroambiguus in Respiratory Function

PurposeThe study aims to elucidate the role and contribution of nucleus retroambiguus (NRA) in generating or modulating the respiratory central pattern generator (rCPG). Previously we identified a putative rhythmogenic area 1.5 mm caudal to obex in the rat in‐situ perfused brainstem‐spinal cord preparation. Histological analysis indicated the putative rhythmogenic cell population resides at the level of the pyramidal decussation, therefore likely to overlap with the NRA. We investigated possible involvement of NRA in rhythmogenesis anatomically and functionally.MethodsAnatomical tracing: Anterograde neuronal tracer, biotinylated dextran amine, was injected into the NRA. Respiratory areas which contained labelled terminal fields received the retrograde tracer Fast Blue in subsequent experiments. Functional testing: Glutamate microinjections were made in a grid formation across the NRA region in the rat in‐situ perfused brainstem spinal cord preparation; with whole nerve recordings taken from phrenic and vagus nerves.ResultsAnterograde and retrograde tracing revealed anatomical connectivity between the NRA and respiratory nuclei, including the pre‐Bötzinger complex, parafacial respiratory group and Kölliker Fuse nucleus. Glutamate mapping in‐situ revealed two distinct responses to pharmacological stimulation. A tachypneic response was seen predominantly overlapping the NRA whilst a breath hold response was seen more frequently in the periphery, dorsal and lateral to the NRA.ConclusionAnatomical evidence shows that NRA can drive respiratory modulation via ascending projections to the rCPG. Therefore the NRA is not just a bulbospinal relay, but rather an integral part of the rCPG.Funding: Melbourne University

Ascending projections of the nucleus retroambiguus target key areas of the ponto‐medullary respiratory network (714.1)

Purpose: We aim to elucidate the role and contribution of the nucleus retroambiguus (NRA) in the generation of respiratory rhythm. We previously identified a putative rhythmogenic area 1.5 mm caudal to obex, destruction of which abolished respiratory rhythm in the in‐situ perfused brainstem‐spinal cord preparation of rat. Histological analysis indicated that the putative rhythmogenic cell population resides at the planar level of the pyramidal decussation, therefore likely to overlap with the NRA. We investigated possible involvement of the NRA in rhythm generation by mapping its neuronal connectivity in rat. Methods: Anterograde neuronal tracer, biotinylated dextran amine, was injected into the NRA. Respiratory areas in which labelled terminal fields were seen were injected with retrograde tracer Fast Blue in subsequent experiments. Results: Anterograde tracing from the NRA revealed dense clusters of labelled nerve terminals within important respiratory nuclei including the pre‐Bötzinger complex (PBC), parafacial respiratory group (pFRG) and Kölliker Fuse nucleus (KF). Injection of retrograde tracer into PBC, KF and pFRG show clusters of retrogradely labelled cell bodies in the NRA. Conclusion: Our data provides the anatomical framework for a rhythmogenic role of the NRA within the respiratory network.Grant Funding Source: ARC

Insufficiency of serotoninergic system at the prenatal period disturbs formation of nucleus retroambiguus in rats

Insufficiency of serotoninergic system at the prenatal period disturbs formation of nucleus retroambiguus in rats

Chapter 20 - The Periaqueductal Gray Controls Brainstem Emotional Motor Systems Including Respiration

Respiration is a motor system essential for the survival of the individual and of the species. Because of its vital significance, studies on respiration often assume that breathing takes place independent of other motor systems. However, motor systems generating vocalization, coughing, sneezing, vomiting, as well as parturition, ejaculation, and defecation encompass abdominal pressure control, which involves changes in the respiratory pattern. The mesencephalic periaqueductal gray (PAG) controls all these motor systems. It determines the level setting of the whole body by means of its very strong projections to the ventromedial medullary tegmentum, but it also controls the cell groups that generate vocalization, coughing, sneezing, vomiting, as well as respiration. For this control, the PAG maintains very strong connections with the nucleus retroambiguus, which enables it to control abdominal and intrathoracic pressure. In this same context, the PAG also runs the pelvic organs, bladder, uterus, prostate, seminal vesicles, and the distal colon and rectum via its projections to the pelvic organ stimulating center and the pelvic floor stimulating center. These cell groups, via long descending projections, have direct control of the parasympathetic motoneurons in the sacral cord as well as of the somatic motoneurons in the nucleus of Onuf, innervating the pelvic floor. Respiration, therefore, is not a motor system that functions by itself, but is strongly regulated by the same systems that also control the other motor output systems.