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.

The midbrain periaqueductal gray as an integrative and interoceptive neural structure for breathing.

The periaqueductal gray (PAG) plays a critical role in autonomic function and behavioural responses to threatening stimuli. Recent evidence has revealed the PAG's potential involvement in the perception of breathlessness, a highly threatening respiratory symptom. In this review, we outline the current evidence in animals and humans on the role of the PAG in respiratory control and in the perception of breathlessness. While recent work has unveiled dissociable brain activity within the lateral PAG during perception of breathlessness and ventrolateral PAG during conditioned anticipation in healthy humans, this is yet to be translated into diseases dominated by breathlessness symptomology, such as chronic obstructive pulmonary disease. Understanding how the sub-structures of the PAG differentially interact with interoceptive brain networks involved in the perception of breathlessness will help towards understanding discordant symptomology, and may reveal treatment targets for those debilitated by chronic and pervasive breathlessness.

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.

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.

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.

The physiological significance of postinspiration in respiratory control.

The term postinspiration is commonly used in the scientific literature concerned with neural generation and the control of breathing movements. Because postinspiration belongs functionally to the mechanical act of expiration, the physiological significance of postinspiration as a distinct phase of the breathing cycle is often underappreciated. The present review will give an overview of the physiological significance of postinspiratory motor activity in laryngeal adductor (constrictor) muscles and the crural diaphragm. The functional importance of postinspiratory motor activity is discussed for the eupneic respiratory cycle, and for various protective respiratory reflex mediations (e.g., sneeze, cough, and breath-hold). Also, the implications of recruited postinspiratory activity during nonventilatory behaviors such as vocalization, swallowing, or vomiting are underpinned. Finally, we describe the impact of absence or malfunction of postinspiratory motor function in neurological diseases.

The midbrain periaqueductal gray changes the eupneic respiratory rhythm into a breathing pattern necessary for survival of the individual and of the species.

Modulation of respiration is a prerequisite for survival of the individual and of the species. For example, respiration has to be adjusted in case of speech, strenuous exercise, laughing, crying, or sudden escape from danger. Respiratory centers in pons and medulla generate the basic respiratory rhythm or eupnea, but they cannot modulate breathing in the context of emotional challenges, for which they need input from higher brain centers. In simple terms, the prefrontal cortex integrates visual, auditory, olfactory, and somatosensory information and informs subcortical structures such as amygdala, hypothalamus, and finally the midbrain periaqueductal gray (PAG) about the results. The PAG, in turn, generates the final motor output for basic survival, such as setting the level of all cells in the brain and spinal cord. Best known in this framework is determining the level of pain perception. The PAG also controls heart rate, blood pressure, micturition, sexual behavior, vocalization, and many other basic motor output systems. Within this context, the PAG also changes the eupneic respiratory rhythm into a breathing pattern necessary for basic survival. This review examines the latest developments regarding of how the PAG controls respiration.

Stimulation of the midbrain periaqueductal gray modulates preinspiratory neurons in the ventrolateral medulla in the rat in vivo.

The midbrain periaqueductal gray (PAG) is involved in many basic survival behaviors that affect respiration. We hypothesized that the PAG promotes these behaviors by changing the firing of preinspiratory (pre-I) neurons in the pre-Bötzinger complex, a cell group thought to be important in generating respiratory rhythm. We tested this hypothesis by recording single unit activity of pre-Bötzinger pre-I neurons during stimulation in different parts of the PAG. Stimulation in the dorsal PAG increased the firing of pre-I neurons, resulting in tachypnea. Stimulation in the medial part of the lateral PAG converted the pre-I neurons into inspiratory phase-spanning cells, resulting in inspiratory apneusis. Stimulation in the lateral part of the lateral PAG generated an early onset of the pre-I neuronal discharge, which continued throughout the inspiratory phase, while at the same time attenuating diaphragm contraction. Stimulation in the ventral part of the lateral PAG induced tachypnea but inhibited pre-I cell firing, whereas stimulation in the ventrolateral PAG inhibited not only pre-I cells but also the diaphragm, leading to apnea. These findings show that PAG stimulation changes the activity of the pre-Bötzinger pre-I neurons. These changes are in line with the different behaviors generated by the PAG, such as the dorsal PAG generating avoidance behavior, the lateral PAG generating fight and flight, and the ventrolateral PAG generating freezing and immobility.

Descending control of the respiratory neuronal network by the midbrain periaqueductal grey in the rat in vivo.

Emotional reactions such as vocalization take place during expiration, and thus expression of emotional behaviour requires a switch from inspiration to expiration. I investigated how the midbrain periaqueductal grey (PAG), a known behavioural modulator of breathing, influences the inspiratory-to-expiratory phase transition. Contemporary models propose that late inspiratory (late-I) and post-inspiratory (post-I) neurones found in the medulla, which are active during the inspiratory-to-expiratory phase transition are involved in converting inspiration to expiration. I examined the effect of excitatory amino acid (d,l-homocysteic acid; DLH) stimulation of the PAG on the discharge function of late-I and post-I neurones. The data show a topographical organization of DLH-induced late-I and post-I neuronal modulation within the PAG. Dorsal PAG stimulation induced tachypnoea and caused excitation of both the late-I and post-I neurones. Lateral PAG induced inspiratory prolongation and caused an excitation of late-I neurones but inhibition of post-I neurones. Ventrolateral PAG induced expiratory prolongation and caused a persistent activation of post-I neurones. As well, PAG stimulation modulated both the late-I and post-I cells for least two-three breaths even prior to the change in respiratory motor pattern. This indicates that the PAG influences the late-I and post-I cells independent of pulmonary or other sensory afferent feedback. I conclude that the PAG modulates the activity of the medullary late-I and post-I neurones, and this modulation contributes to the conversion of eupnoea into a behavioural breathing pattern.

The nucleus retroambiguus as possible site for inspiratory rhythm generation caudal to obex.

We investigated whether spinalized animals can produce inspiratory rhythm. We recorded spinal inspiratory phrenic (PNA) and cranial inspiratory hypoglossal (HNA) nerve activity in the perfused brainstem preparation of rat. Complete transverse transections were performed at 1.5 (pyramidal decussation) or 2mm (first cervical spinal segment) caudal to obex. Excitatory drive was enhanced by either extracellular potassium, hypercapnia or by stimulating arterial chemoreceptors. Caudal transections immediately eliminated descending network drive for PNA, while the cranial inspiratory HNA remained unaffected. After transection, PNA bursting remained sporadic even during enhanced excitatory drive. This implies, cervical spinal circuits lack intrinsic rhythmogenic capacity. Rostral transections also abolished PNA immediately. However, HNA also progressively lost its amplitude and rhythm. Chemoreceptor activation only triggered tonic, non-rhythmic HNA. Thus the integrity of ponto-medullary circuitry was maintained. Our results suggest that an area overlapping the caudal nucleus retroambiguus provides critical ascending input to the ponto-medullary respiratory network for inspiratory rhythm generation.

Midbrain and medullary control of postinspiratory activity of the crural and costal diaphragm in vivo.

Studies on brain stem respiratory neurons suggest that eupnea consists of three phases: inspiration, postinspiration, and expiration. However, it is not well understood how postinspiration is organized in the diaphragm, i.e., whether postinspiration differs in the crural and costal segments of the diaphragm and what the influence is of postinspiratory neurons on diaphragm function during eupnea. In this in vivo study we investigated the postinspiratory activity of the two diaphragm segments during eupnea and the changes in diaphragm function following modulation of eupnea. Postinspiratory neurons in the medulla were stereotaxically localized extracellularly and neurochemically stimulated. We used three types of preparations: precollicularly decerebrated unanesthetized cats and rats and anesthetized rats. In all preparations, during eupnea, postinspiratory activity was found in the crural but not in the costal diaphragm. When eupnea was discontinued in decerebrate cats in which stimulation in the nucleus retroambiguus induced activation of laryngeal or abdominal muscles, all postinspiratory activity in the crural diaphragm was abolished. In decerebrate rats, stimulation of the midbrain periaqueductal gray abolished postinspiration in the crural diaphragm but induced activation in the costal diaphragm. In anesthetized rats, stimulation of medullary postinspiratory neurons abolished the postinspiratory activity of the crural diaphragm. Vagal nerve stimulation in these rats increased the intensity of postinspiratory neuronal discharge in the solitary nucleus, leading to decreased activity of the crural diaphragm. These data demonstrate that three-phase breathing in the crural diaphragm during eupnea exists in vivo and that postinspiratory neurons have an inhibitory effect on crural diaphragm function.

The role of serotonin in respiratory function and dysfunction.

Serotonin (5-HT) is a neuromodulator-transmitter influencing global brain function. Past and present findings illustrate a prominent role for 5-HT in the modulation of ponto-medullary autonomic circuits. 5-HT is also involved in the control of neurotrophic processes during pre- and postnatal development of neural circuits. The functional implications of 5-HT are particularly illustrated in the alterations to the serotonergic system, as seen in a wide range of neurological disorders. This article reviews the role of 5-HT in the development and control of respiratory networks in the ponto-medullary brainstem. The review further examines the role of 5-HT in breathing disorders occurring at different stages of life, in particular, the neonatal neurodevelopmental diseases such as Rett, sudden infant death and Prader-Willi syndromes, adult diseases such as sleep apnoea and mental illness linked to neurodegeneration.

Periaqueductal gray control of breathing.

Change of the basic respiratory rhythm (eupnea) is a pre-requisite for survival. For example, sudden escape from danger needs rapid shallow breathing, strenuous exercise requires tachypnea for sufficient supply of oxygen and a strong anxiety reaction necessitates gasping. Also for vocalization (and for speech in humans) an important mechanism for survival, respiration has to be changed. The caudal brainstem premotor respiratory centers need input from higher brain centers in order to change respiration according to the surrounding circumstances. One of the most important of such a higher brain centers is the midbrain periaqueductal gray (PAG). The PAG co-ordinates motor output, including respiratory changes based on input from limbic, prefrontal and anterior cingulate cortex regions. These areas integrate visual, auditory and somatosensory information in the context of basic survival mechanisms and relay the result to the PAG, which has access to respiratory control centers in the caudal brainstem. Through these pathways the PAG can change eupneic respiratory rhythm into the behavior necessary for that specific situation. We present data obtained from the cat and propose a functional framework for the breathing control pathways.

The nucleus retroambiguus control of respiration.

The role of the nucleus retroambiguus (NRA) in the context of respiration control has been subject of debate for considerable time. To solve this problem, we chemically (using d, l-homocysteic acid) stimulated the NRA in unanesthetized precollicularly decerebrated cats and studied the respiratory effect via simultaneous measurement of tracheal pressure and electromyograms of diaphragm, internal intercostal (IIC), cricothyroid (CT), and external oblique abdominal (EO) muscles. NRA-stimulation 0-1 mm caudal to the obex resulted in recruitment of IIC muscle and reduction in respiratory frequency. NRA-stimulation 1-3 mm caudal to the obex produced vocalization along with CT activation and slight increase in tracheal pressure, but no change in respiratory frequency. NRA-stimulation 3-5 mm caudal to the obex produced CT muscle activation and an increase in respiratory frequency, but no vocalization. NRA-stimulation 5-8 mm caudal to the obex produced EO muscle activation and reduction in respiratory frequency. A change to the inspiratory effort was never observed, regardless of which NRA part was stimulated. The results demonstrate that NRA does not control eupneic inspiration but consists of topographically separate groups of premotor interneurons each producing detailed motor actions. These motor activities have in common that they require changes to eupneic breathing. Different combination of activation of these premotor neurons determines the final outcome, e.g., vocalization, vomiting, coughing, sneezing, mating posture, or child delivery. Higher brainstem regions such as the midbrain periaqueductal gray (PAG) decides which combination of NRA neurons are excited. In simple terms, the NRA is the piano, the PAG one of the piano players.

The midbrain periaqueductal gray control of respiration.

The midbrain periaqueductal gray (PAG) organizes basic survival behavior, which includes respiration. How the PAG controls respiration is not known. We studied the PAG control of respiration by injecting D,L-homocysteic acid in the PAG in unanesthetized precollicularly decerebrated cats. Injections in different parts of the PAG caused different respiratory effects. Stimulation in the dorsomedial PAG induced slow and deep breathing and dyspnea. Stimulation in the dorsolateral PAG resulted in active breathing and tachypnea consistent with the respiratory changes during fright and flight. Stimulation in the medial part of lateral PAG caused inspiratory apneusis. Stimulation in lateral parts of the lateral and ventrolateral PAG produced respiratory changes associated with vocalization (mews, alternating mews and hisses, or hisses). D,L-homocysteic acid injections in the caudal ventrolateral PAG induced irregular breathing. These results demonstrate that the PAG exerts a strong influence on respiration, suggesting that it serves as the behavioral modulator of breathing.

Ventilation induced apnea and its effect on dorsal brainstem inspiratory neurones in the rat.

The purpose of this study was to examine the effect of mechanical ventilation (MV) on inherent breathing and on dorsal brainstem nucleus tractus solitarius (NTS) respiratory cell function. In pentobarbitone-anaesthetised rats, application of MV at combined high frequencies and volumes (representing threshold levels) produced apnea. The apnea persisted as long as MV was maintained at or above the threshold frequency and volume. Following removal of MV, inherent breathing did not resume immediately, with the diaphragm exhibiting post-mechanical ventilation apnea. The fall in arterial P(CO2) (Pa(CO2)) levels evoked by MV-engendered hyperventilation was shown not to be the trigger for initiation of apnea. MV-induced apnea was immediately reversed by bilateral vagotomy. Further, MV-induced apnea could not be evoked in bilaterally vagotomized animals suggesting that vagal feedback is the critical pathway for its initiation. NTS inspiratory neurones were inhibited during both MV-induced apnea and post-mechanical ventilation apnea, implying the involvement of central neural mechanisms in mediating this effect.

Identification of different types of respiratory neurones in the dorsal brainstem nucleus tractus solitarius of the rat.

In Nembutal anaesthetised, spontaneously breathing rats, stereotaxic mapping of the nucleus tractus solitarius (NTS) for respiratory neuronal activity was undertaken. Eight different types of respiratory cells were found between 0.25 and 1.5 mm lateral to midline, extending 0.5 mm caudal to 1.5 mm rostral to obex, and 0.4-1.5 mm below the dorsal surface. A study of the respiratory motor (diaphragm EMG) and neuronal responses to excitatory amino acid (EAA) stimulation of the NTS areas was undertaken. Electrical stimulation of the vagus nerve was employed to study the NTS cellular responses to activation of pulmonary afferents. The effects of chemical activation of the midbrain periaqueductal grey (PAG) on NTS respiratory neuronal activity were investigated. EAA microinjections into the ventrolateral NTS rostral to the obex resulted in an increase in respiratory motor frequency along with increases to inspiratory cell discharge, whilst microinjections into the medial NTS caudal to the obex caused respiratory depression. EAA stimulation of calamus scriptorius produced apnea. NTS inspiratory neurones were inhibited following stimulation of ipsilateral vagus nerve, suggesting their involvement in the Hering-Breuer reflex pathway. PAG stimulation caused excitation of the NTS inspiratory cells indicating the presence of an excitatory respiratory pathway between the two nuclei. Following beta-adrenergic antagonist pre-treatment of ventrolateral NTS, EAA microinjections into PAG did not evoke a cardiorespiratory effect. Based on the various findings the role of NTS in organising respiration in the rat is discussed.