Paraventricular oxytocin neurons are involved in neural modulation of breathing.

In this study, we determined the projections of oxytocin-containing neurons of the paraventricular nucleus (PVN) to phrenic nuclei and to the rostral ventrolateral medullary (RVLM) region, which is known to be involved in respiratory rhythm generation. Studies were also designed to determine oxytocin-receptor expression within the RVLM and the physiological effects of their activation on respiratory drive and arterial blood pressure. Oxytocin immunohistochemistry combined with cholera toxin B, a retrograde tracer, showed that a subpopulation of oxytocin-containing parvocellular neurons in the dorsal and medial ventral regions of the PVN projects to phrenic nuclei. Similarly, a subpopulation of pseudorabies virus-labeled neurons in the PVN coexpressed oxytocin after injection of pseudorabies virus, a transynaptic retrograde marker, into the costal region of the diaphragm. A subpopulation of oxytocin expressing neurons was also found to project to the RVLM. Activation of this site by microinjection of oxytocin into the RVLM (0.2 nmol/200 nl) significantly increased diaphragm electromyographic activity and frequency discharge (P < 0.05). In addition, oxytocin increased blood pressure and heart rate (P < 0.05). These data indicate that oxytocin participates in the regulation of respiratory and cardiovascular activity, partly via projections to the RVLM and phrenic nuclei.

Monoaminergic neurons, chemosensation and arousal.

In recent years, immense progress has been made in understanding central chemosensitivity at the cellular and functional levels. Combining molecular biological techniques (early gene expression as an index of cell activation) with neurotransmitter immunohistochemistry, new information has been generated related to neurochemical coding in chemosensory cells. We found that CO(2) exposure leads to activation of discrete cell groups along the neuraxis, including subsets of cells belonging to monoaminergic cells, noradrenaline-, serotonin-, and histamine-containing neurons. In part, they may play a modulatory role in the respiratory response to hypercapnia that could be related to their behavioral state control function. Activation of monoaminergic neurons by an increase in CO(2)/H(+) could facilitate respiratory related motor discharge, particularly activity of upper airway dilating muscles. In addition, these neurons coordinate sympathetic and parasympathetic tone to visceral organs, and participate in adjustments of blood flow with the level of motor activity. Any deficit in CO(2) chemosensitivity of a network composed of inter-related monoaminergic nuclei might lead to disfacilitation of motor outputs and to failure of neuroendocrine and homeostatic responses to life-threatening challenges (e.g. asphyxia) during sleep.

Differential cardiorespiratory control elicited by activation of ventral medullary sites in mice.

We studied the respiratory and blood pressure responses to chemical stimulation of two regions of the ventral brainstem in mice: the rostral and caudal ventrolateral medulla (RVLM and CVLM, respectively). Stimulation of the RVLM by microinjections of the excitatory amino acid L-glutamate induced increases in diaphragm activity and breathing frequency, elevation of blood pressure (BP), and a slight increase in heart rate (HR). However, activation of the CVLM induced a decrease in breathing frequency, mainly due to prolongation of expiratory time (TE), and hypotension associated with a slight slowing of HR. Because adrenergic mechanisms are known to participate in the control of respiratory timing, we examined the role of alpha(2)-adrenergic receptors in the RVLM region in mediating these inhibitory effects. The findings demonstrated that blockade of the alpha(2)-adrenergic receptors within the RVLM by prior microinjection of SKF-86466 (an alpha(2)-adrenergic receptor blocker) significantly reduced changes in TE induced by CVLM stimulation but had little effect on BP responses. These results indicate that, in mice, activation of the RVLM increases respiratory drive associated with an elevation of BP, but stimulation of CVLM induces prolongation of TE via an alpha(2)-adrenergic signal transduction pathway.

alpha(2)-adrenergic receptors are not required for central anti-hypertensive action of moxonidine in mice.

In the mouse medulla oblongata, we characterized binding properties and functional responses of two recognition sites for imidazoline compounds: I(1)-imidazoline and alpha(2)-adrenergic receptors. The mouse medulla expresses a higher density of I(1)-receptors than in the rat, whereas alpha(2)-receptor densities were similar between the two species. In anesthetized, ventilated and paralyzed mice, we tested the hypotensive actions of the I(1)/alpha(2) agonist moxonidine, determined its central site of its actions, and the relative roles of I(1) and alpha(2)-receptors. Experiments were performed in C(57)Bl(6) wild type and alpha(2A)-adrenergic receptor deficient mice. In both types of mice, neuronal activation within the rostral ventrolateral medulla (RVLM) region by glutamate microinjection elicited increases in arterial pressure. Moxonidine (0.5 nmol/site/10 nl) microinjected bilaterally into this vasopressor region decreased arterial pressure by 30% and heart rate by 11% in wild type mice. Efaroxan, the I(1)/alpha(2) antagonist (0.4 nmol) when microinjected into the RVLM elevated blood pressure itself and abolished the action of moxonidine, whereas alpha(2)-blockade with SK&F 86466 had no significant effect on blood pressure and did not attenuate moxonidine's effect. To more definitively test the role of alpha(2)-adrenergic receptors in the action of moxonidine, moxonidine was microinjected into the RVLM of alpha(2A)-adrenergic deficient mice. The decreases in arterial pressure were nearly identical to those of wild type mice, whereas bradycardia was attenuated. Thus, in the mouse moxonidine acts within the RVLM region to lower arterial pressure mainly through the I(1)-imidazoline receptor independent of alpha(2)-adrenergic receptors.

Disruption of the murine nuclear factor I-A gene (Nfia) results in perinatal lethality, hydrocephalus, and agenesis of the corpus callosum.

The phylogenetically conserved nuclear factor I (NFI) family of transcription/replication proteins is essential both for adenoviral DNA replication and for the transcription of many cellular genes. We showed previously that the four murine NFI genes (Nfia, Nfib, Nfic, and Nfix) are expressed in unique but overlapping patterns during mouse development and in adult tissues. Here we show that disruption of the Nfia gene causes perinatal lethality, with >95% of homozygous Nfia(-/-) animals dying within 2 weeks after birth. Newborn Nfia(-/-) animals lack a corpus callosum and show ventricular dilation indicating early hydrocephalus. Rare surviving homozygous Nfia(-/-) mice lack a corpus callosum, show severe communicating hydrocephalus, a full-axial tremor indicative of neurological defects, male-sterility, low female fertility, but near normal life spans. These findings indicate that while the Nfia gene appears nonessential for cell viability and DNA replication in embryonic stem cells and fibroblasts, loss of Nfia function causes severe developmental defects. This finding of an NFI gene required for a developmental process suggests that the four NFI genes may have distinct roles in vertebrate development.

Respiratory effects of kynurenic acid microinjected into the ventromedullary surface of the rat

Several studies demonstrate that, within the ventral medullary surface (VMS), excitatory amino acids are necessary components of the neural circuits involved in the tonic and reflex control of respiration and circulation. In the present study we investigated the cardiorespiratory effects of unilateral microinjections of the broad spectrum glutamate antagonist kynurenic acid (2 nmol/200 nl) along the VMS of urethane-anesthetized rats. Within the VMS only one region was responsive to this drug. This area includes most of the intermediate respiratory area, partially overlapping the rostral ventrolateral medulla (IA/RVL). When microinjected into the IA/RVL, kynurenic acid produced a respiratory depression, without changes in mean arterial pressure or heart rate. The respiratory depression observed was characterized by a decrease in ventilation, tidal volume and mean inspiratory flow and an increase in respiratory frequency. Therefore, the observed respiratory depression was entirely due to a reduction in the inspiratory drive. Microinjections of vehicle (200 nl of saline) into this area produced no significant changes in breathing pattern, blood pressure or heart rate. Respiratory depression in response to the blockade of glutamatergic receptors inside the rostral VMS suggests that neurons at this site have an endogenous glutamatergic input controlling the respiratory cycle duration and the inspiratory drive transmission.

Respiratory effects of kynurenic acid microinjected into the ventromedullary surface of the rat.

Several studies demonstrate that, within the ventral medullary surface (VMS), excitatory amino acids are necessary components of the neural circuits involved in the tonic and reflex control of respiration and circulation. In the present study we investigated the cardiorespiratory effects of unilateral microinjections of the broad spectrum glutamate antagonist kynurenic acid (2 nmol/200 nl) along the VMS of urethane-anesthetized rats. Within the VMS only one region was responsive to this drug. This area includes most of the intermediate respiratory area, partially overlapping the rostral ventrolateral medulla (IA/RVL). When microinjected into the IA/RVL, kynurenic acid produced a respiratory depression, without changes in mean arterial pressure or heart rate. The respiratory depression observed was characterized by a decrease in ventilation, tidal volume and mean inspiratory flow and an increase in respiratory frequency. Therefore, the observed respiratory depression was entirely due to a reduction in the inspiratory drive. Microinjections of vehicle (200 nl of saline) into this area produced no significant changes in breathing pattern, blood pressure or heart rate. Respiratory depression in response to the blockade of glutamatergic receptors inside the rostral VMS suggests that neurons at this site have an endogenous glutamatergic input controlling the respiratory cycle duration and the inspiratory drive transmission.

Cardiorespiratory effects of L-glutamate microinjected into the rat ventral medulla.

We investigated the cardiorespiratory effects elicited by microinjections of L-glutamate (L-glu, 25 nmol, 200 nl) at various sites in the ventral medulla (VMS) of urethane-anesthetized rats. The results demonstrated that regions responsive to the drug are located along a column in the VMS extending from the VI cranial nerve to the first cervical nerve in the caudal medulla. Within this column three breathing patterns were elicited from four distinct areas. In the most rostral and caudal portion of this hypothetical column, the breathing patterns observed in response to L-glu were similar and characterized by increases in minute ventilation, tidal volume, inspiratory drive, respiratory frequency, mean arterial blood pressure (MAP) and heart rate (HR). In the regions located between the areas described above two different breathing patterns were obtained without significant changes in MAP or HR. These patterns were characterized by decreases and increases in the respiratory indices analyzed, with the exception of respiratory frequency, which decreased in both regions. These results suggest that within the VMS discrete areas may act as functional units modulating cardiorespiratory responses while in others these functions are spatially segregated.

Cardiorespiratory alterations produced by pharmacological agents applied to the ventrolateral medulla (intermediate area) of the cat. Effect of exercise condition on glycine-induced inhibition.

1. To study the action of the intermediate area (IA), coextensive with the rostral ventrolateral medulla, on the neurophysiological mechanisms involved in the regulation of respiration, in terms of inspiratory drive and respiratory timing, cats were submitted to topical application of sodium pentobarbital (30 mg/ml), leptazol (200 mg/ml), glutamate (50 mg/ml) and glycine (100 and 50 mg/ml) to the IA. The effects of electrically induced exercise on the ventilatory response and oxygen uptake (VO2) obtained by topical application of glycine (50 mg/ml) to the IA were also studied. 2. Leptazol reduced minute ventilation (VE) and inspiratory drive (VT/TI) and changed the timing mechanism. Glutamate only increased tidal volume (VT), VE and VT/TI. Arterial blood pressure (AP) increased and heart rate (HR) did not change with either drug. 3. Sodium pentobarbital reduced VT and changed the timing mechanism. Glycine only reduced VE, VT and VT/TI. AP decreased and HR did not change with either drug. 4. The depressor effects of glycine on respiratory pattern, VO2 and CO2 production (VCO2) tended to be attenuated by exercise. 5. The fall in AP due to glycine application did not differ between resting and exercise conditions. 6. Our results indicate that at least two different nervous structures are involved in the IA: one responsible for the respiratory drive and sensitive to glycine and glutamate, and the other responsible for the regulation of the timing mechanism and sensitive to sodium pentobarbital and leptazol.