Uncoupling of upper airway motor activity from phrenic bursting by positive end-expired pressure in the rat.

Phasic bursting in the hypoglossal nerve can be uncoupled from phrenic bursting by application of positive end-expired pressure (PEEP). We wished to determine whether similar uncoupling can also be induced in other respiratory-modulated upper airway (UAW) motor outputs. Discharge of the facial, hypoglossal, superior laryngeal, recurrent laryngeal, and phrenic nerves was recorded in anesthetized, ventilated rats during stepwise changes in PEEP with a normocapnic, hyperoxic background. Application of 3- to 6-cmH(2)O PEEP caused the onset inspiratory (I) UAW nerve bursting to precede the phrenic burst but did not uncouple bursting. In contrast, application of 9- to 12-cmH(2)O PEEP uncoupled UAW neurograms such that rhythmic bursting occurred during periods of phrenic quiescence. Single-fiber recording experiments were conducted to determine whether a specific population of UAW motoneurons is recruited during uncoupled bursting. The data indicate that expiratory-inspiratory (EI) motoneurons remained active, while I motoneurons did not fire during uncoupled UAW bursting. Finally, we examined the relationship between motoneuron discharge rate and PEEP during coupled UAW and phrenic bursting. EI discharge rate was linearly related to PEEP during preinspiration, but showed no relationship to PEEP during inspiration. Our results demonstrate that multiple UAW motor outputs can be uncoupled from phrenic bursting, and this response is associated with bursting of EI nerve fibers. The relationship between PEEP and EI motoneuron discharge rate differs during preinspiratory and I periods; this may indicate that bursting during these phases of the respiratory cycle is controlled by distinct neuronal outputs.     

Pulmonary C-fiber activation enhances respiratory-related activities of the recurrent laryngeal nerve in rats

The purpose of the current study was to characterize the response of the recurrent laryngeal nerve (RLN) to pulmonary C-fiber activation. Male rats of Wistar strain were anesthetized by urethane (1.2 g/kg, i.p.). Tracheostomy was performed. Catheter was inserted into the femoral artery and vein. Additional catheter was placed near the entrance of the right atrium via the right jugular vein. The animal was then paralyzed with gallamine triethiodide, ventilated and maintained at normocapnia in hyperoxia. Activities of the phrenic (PNA) and recurrent laryngeal nerves (RLNA) were monitored simultaneously. Two experimental protocols were completed. In the first experiment, various doses of capsaicin were delivered into the right atrium to activate pulmonary C-fibers with vagal intact. Low dose of capsaicin (1.25 microg/kg) produced apnea, a decrease in amplitude of PNA, an enhancement of RLNA during apnea and recovery from apnea, hypotension, and bradycardia. High dose of capsaicin (5 and 20 microg/kg) evoked the same tendency of response for both nerves and biphasic changes in blood pressure. Dose dependency was only seen in the period of apnea but not observable in nerve amplitudes. After bilateral vagotomy, low dose of capsaicin produced an increase in PNA without apnea, no significant change in RLNA, and hypertension. These results suggest that activation of vagal and nonvagal C-fibers could produce different reflex effects on cardiopulmonary functions. The reflex responses evoked by these two types of afferents might play defensive and protective roles in the airways and lungs.     

Reflex mechanisms for changes in renal nerve activity during positive end-expiratory pressure

This study was designed to investigate the interaction between carotid sinus baroreceptors and cardiopulmonary receptors in the reflex control of renal nerve activity (RNA) during positive end-expiratory pressure (PEEP) in anesthetized dogs. PEEP at two different levels (10 and 20 cmH2O) was applied to the following groups: animals with neuraxis intact (I group, n = 12); vagal and aortic nerve denervated animals with carotid sinus nerves intact (V group, n = 6); carotid sinus denervated animals with vagal and aortic nerves intact (SD group, n = 6); and carotid sinus denervated animals also having severed vagal and aortic nerves (SAV group, n = 12). Mean blood pressure (MBP), central venous pressure, and mean airway pressure were also simultaneously measured. In the I group, no significant alterations in RNA occurred during PEEP at both levels, even when MBP fell significantly. Although the drop in MBP in the SD group was similar to that in the I group, RNA decreased significantly 10 s after intervention at both PEEP levels, followed by a recovery of RNA toward the control level. In contrast, a significant increase in RNA, which continued until the end of PEEP, appeared in the V group immediately after each intervention. In the SAV group, RNA responses to PEEP, which were observed in the other groups, were abolished. These results provide evidence that during PEEP, renal nerve activity is modified by an interaction between carotid sinus baroreceptors and cardiopulmonary receptors; excitatory effects occur via carotid sinus nerves and inhibitory effects occur via vagal afferents.     

Spontaneous phrenic nerve activity in the exteriorized fetal lamb.

Phrenic nerve activity and tracheal pressure changes were recorded in four exteriorized fetal lambs (120-135 days gestation) from lightly anesthetized ewes to study possible mechanisms involved in the establishment of rhythmical breathing patterns. Two types of spontaneous neural activity were found. The first consisted of high-frequency multiunit bursts (mean duration 820 ms; range 450-2,500 ms) that preceded a gasp. Individual units within these bursts reached peak discharge frequencies as high as 40 impulses/s. The second type of neural activity consisted of single-unit, low-frequency (1-14 impulses/s), irregular background discharges lasting up to several seconds without changes in tracheal pressure. Occasionally, higher frequency bursts of single-unit activity were detected that were also unassociated with tracheal pressure changes. The data indicate that the neural correlate of a fetal gasp includes high-frequency synchronized bursting activity in the phrenic nerve. In addition, background phrenic activity can be detected in the exteriorized fetal lamb that reflects central nervous activity in the absence of tracheal pressure changes.     

Efferent activity in the phrenic nerve during startle reflex in chloralose anesthetized cats

Efferent activity was investigated in the phrenic nerve during startle reflex manifesting as somatic nerve discharges (lower intercostal nerves and the nerve endings) in chloralose anesthetized cats. Inhibition (usually of short duration, lasting 23–36 msec) of inspiration activity was found to be the main component of response in the phrenic nerve in the shaping of "low threshold" startle reflex produced by acoustic and tactile stimuli and stimulation of low threshold peripheral afferents. Reflex discharge prevailed amongst the response patterns produced in the phrenic nerve by stimulating high threshold afferents, i.e., early (propriospinal) and late (suprasegmental, arising from stimulating intercostal nerve) or late only (when stimulating the hindlimb nerves). Two patterns of late response could be distinguished, one on inspiration (found in roughly 3 out of 4 experiments) and other on exhalation — the respiratory homologs of somatic startle reflex. Response pattern is described throughout the respiratory cycle. Structure and respiratory modulation of reflex responses produced in the phrenic nerve by stimulating bulbar respiratory structure are also examined. Possible neurophysiological mechanisms underlying phrenic response during the shaping of startle reflex are discussed.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 19, No. 4, pp. 473–482, July–August, 1987.     

Entrainment pattern between sympathetic and phrenic nerve activities in the Sprague-Dawley rat: hypoxia-evoked sympathetic activity during expiration

Sympathetic and respiratory motor activities are entrained centrally. We hypothesize that this coupling may partially underlie changes in sympathetic activity evoked by hypoxia due to activity-dependent changes in the respiratory pattern. Specifically, we tested the hypothesis that sympathetic nerve activity (SNA) expresses a short-term potentiation in activity after hypoxia similar to that expressed in phrenic nerve activity (PNA). Adult male, Sprague-Dawley (Zivic Miller) rats (n = 19) were anesthetized (Equithesin), vagotomized, paralyzed, ventilated, and pneumothoracotomized. We recorded PNA and splanchnic SNA (sSNA) and generated cycle-triggered averages (CTAs) of rectified and integrated sSNA before, during, and after exposures to hypoxia (8% O(2) and 92% N(2) for 45 s). Inspiration (I) and expiration (E) were divided in half, and the average and area of integrated sSNA were calculated and compared at the following time points: before hypoxia, at the peak breathing frequency during hypoxia, immediately before the end of hypoxia, immediately after hypoxia, and 60 s after hypoxia. In our animal model, sSNA bursts consistently followed the I-E phase transition. With hypoxia, sSNA increased in both halves of E, but preferentially in the second rather than the first half of E, and decreased in I. After hypoxia, sSNA decreased abruptly, but the coefficient of variation in respiratory modulation of sSNA was significantly less than that at baseline. The hypoxic-evoked changes in sympathetic activity and respiratory pattern resulted in sSNA in the first half of E being correlated negatively to that in the second half of E (r = -0.65, P < 0.05) and positively to Te (r = 0.40, P < 0.05). Short-term potentiation in sSNA appeared not as an increase in the magnitude of activity but as an increased consistency of its respiratory modulation. By 60 s after hypoxia, the variability in the entrainment pattern had returned to baseline. The preferential recruitment of late expiratory sSNA during hypoxia results from either activation by expiratory-modulated neurons or by non-modulated neurons whose excitatory drive is not gated during late E.     

Control of phrenic nerve activity and blood pressure by the medullary raphe nuclei in cats

Electrical and chemical stimulation methods were used to determine the topographic organization of the medullary raphe nuclei (MRN) in controlling the systemic arterial blood pressure (BP) and phrenic nerve activities (PNA). Decerebrated, unanesthetized and bilateral vagotomized cats were used. Effective points in the MRN were systematically explored with constant current stimulation. We found stimulation of the rostral MRN produced a decrease in PNA amplitude and increase in BP and PNA frequency. Stimulation of the caudal MRN produced increases in BP and the amplitude and frequency of PNA. Microinjection of glutamate solution into the caudal or the rostral MRN points produced qualitatively similar results. Thus, we concluded that the caudal MRN neurons had excitatory connections whereas the rostral MRN neurons had excitatory and inhibitory connections to the cardiovascular preganglionic neurons and the phrenic nerve motoneurons.     

An overview of phrenic nerve and diaphragm muscle development in the perinatal rat.

In this overview, we outline what is known regarding the key developmental stages of phrenic nerve and diaphragm formation in perinatal rats. These developmental events include the following. Cervical axons emerge from the spinal cord during embryonic (E) day 11. At approximately E12.5, phrenic and brachial axons from the cervical segments merge at the brachial plexi. Subsequently, the two populations diverge as phrenic axons continue to grow ventrally toward the diaphragmatic primordium and brachial axons turn laterally to grow into the limb bud. A few pioneer axons extend ahead of the majority of the phrenic axonal population and migrate along a well-defined track toward the primordial diaphragm, which they reach by E13.5. The primordial diaphragmatic muscle arises from the pleuroperitoneal fold, a triangular protrusion of the body wall composed of the fusion of the primordial pleuroperitoneal and pleuropericardial tissues. The phrenic nerve initiates branching within the diaphragm at approximately E14, when myoblasts in the region of contact with the phrenic nerve begin to fuse and form distinct primary myotubes. As the nerve migrates through the various sectors of the diaphragm, myoblasts along the nerve's path begin to fuse and form additional myotubes. The phrenic nerve intramuscular branching and concomitant diaphragmatic myotube formation continue to progress up until E17, at which time the mature pattern of innervation and muscle architecture are approximated. E17 is also the time of the commencement of inspiratory drive transmission to phrenic motoneurons (PMNs) and the arrival of phrenic afferents to the motoneuron pool. During the period spanning from E17 to birth (gestation period of approximately 21 days), there is dramatic change in PMN morphology as the dendritic branching is rearranged into the rostrocaudal bundling characteristic of mature PMNs. This period is also a time of significant changes in PMN passive membrane properties, action-potential characteristics, and firing properties.     

Selective loss of high-frequency oscillations in phrenic and hypoglossal activity in the decerebrate rat during gasping

Respiratory motor outputs contain medium-(MFO) and high-frequency oscillations (HFO) that are much faster than the fundamental breathing rhythm. However, the associated changes in power spectral characteristics of the major respiratory outputs in unanesthetized animals during the transition from normal eupneic breathing to hypoxic gasping have not been well characterized. Experiments were performed on nine unanesthetized, chemo- and barodenervated, decerebrate adult rats, in which asphyxia elicited hyperpnea, followed by apnea and gasping. A gated fast Fourier transform (FFT) analysis and a novel time-frequency representation (TFR) analysis were developed and applied to whole phrenic and to medial branch hypoglossal nerve recordings. Our results revealed one MFO and one HFO peak in the phrenic output during eupnea, where HFO was prominent in the first two-thirds of the burst and MFO was prominent in the latter two-thirds of the burst. The hypoglossal activity contained broadband power distribution with several distinct peaks. During gasping, two high-amplitude MFO peaks were present in phrenic activity, and this state was characterized by a conspicuous loss in HFO power. Hypoglossal activity showed a significant reduction in power and a shift in its distribution toward lower frequencies during gasping. TFR analysis of phrenic activity revealed the increasing importance of an initial low-frequency "start-up" burst that grew in relative intensity as hypoxic conditions persisted. Significant changes in MFO and HFO rhythm generation during the transition from eupnea to gasping presumably reflect a reconfiguration of the respiratory network and/or alterations in signal processing by the circuitry associated with the two motor pools.     

Changes of impulse activity recorded from the rat vagus nerve during short-lasting total cooling

Gradual cooling of anesthetized rats followed by a drop in rectal temperature (RT) increased the frequency of efferent impulses and decreased the frequency of afferent impulses in the vagus nerves. Preliminary short-lasting (5 h) moderate cooling of the animals in a thermochamber to +5°C (RT did not change), or intensive cooling to −20°C (RT dropped to 32°C) changed the response of efferent nerve fibers to cooling of the body. Under these conditions, a drop in RT to 29°C was followed by a significant increase in efferent discharges in the vagus nerve after additional cooling throughout the experiment, while an initial cooling phase (RT was equal to 35-30°C) was followed by some inhibitory effect. At the same time, the changes in the afferent link were different. As in the control, gradual cooling decreased frequency of afferent impulses, although the intensity of the effects was different. The involvement of the vagus nerve system in the maintenance of temperature homeostasis during body cooling has been discussed.     

Recording of blood pressure,heart rate and aortic nerve activity during parabolic flight in the rat via radio-telemetry

Exposure to microgravity induces cardiovascular deconditioning characterized by orthostatic hypotension when astronauts return to the earth. In order to understand the mechanism of cardiovascular deconditioning, it is necessary to clarify the changes in hemodynamics and the cardiovascular regulation system over the period of space flight. The telemetry system applied to freely moving animals will be a useful and appropriate technique for this kind of long term study of the cardiovascular system in the conscious animal during space flight. The purpose of the present study is twofold: firstly, to observe the detailed changes of arterial pressure and heart rate (HR) during microgravity elicited by the parabolic flight in order to study the acute effect of microgravity exposure on the cardiovascular system; and secondly, to test the feasibility of the telemetry system for recording blood pressure, HR and autonomic nervous activities continuously during space flight.     

Medullary lateral tegmental field neurons influence the timing and pattern of phrenic nerve activity in cats.

In an effort to characterize the role of the medullary lateral tegmental field (LTF) in regulating respiration, we tested the effects of selective blockade of excitatory (EAA) and inhibitory amino acid (IAA) receptors in this region on phrenic nerve activity (PNA) of vagus-intact and vagotomized cats anesthetized with dial-urethane. We found distinct patterns of changes in central respiratory rate, duration of inspiratory and expiratory phases of PNA (Ti and Te, respectively), and I-burst amplitude after selective blockade of EAA and IAA receptors in the LTF. First, blockade of N-methyl-D-aspartate (NMDA) receptors significantly (P < 0.05) decreased central respiratory rate primarily by increasing Ti but did not alter I-burst amplitude. Second, blockade of non-NMDA receptors significantly reduced I-burst amplitude without affecting central respiratory rate. Third, blockade of GABAA receptors significantly decreased central respiratory rate by increasing Te and significantly reduced I-burst amplitude. Fourth, blockade of glycine receptors significantly decreased central respiratory rate by causing proportional increases in Ti and Te and significantly reduced I-burst amplitude. These changes in PNA were markedly different from those produced by blockade of EAA or IAA receptors in the pre-B?tzinger complex. We propose that a proper balance of excitatory and inhibitory inputs to several functionally distinct pools of LTF neurons is essential for maintaining the normal pattern of PNA in anesthetized cats.     

Muscle sympathetic nerve activity during lower body negative pressure is accentuated in heat-stressed humans.

The purpose of this project was to test the hypothesis that increases in muscle sympathetic nerve activity (MSNA) during an orthostatic challenge is attenuated in heat-stressed individuals. To accomplish this objective, MSNA was measured during graded lower body negative pressure (LBNP) in nine subjects under normothermic and heat-stressed conditions. Progressive LBNP was applied at -3, -6, -9, -12, -15, -18, -21, and -40 mmHg for 2 min per stage. Whole body heating caused significant increases in sublingual temperature, skin blood flow, sweat rate, heart rate, and MSNA (all P < 0.05) but not in mean arterial blood pressure (P > 0.05). Progressive LBNP induced significant increases in MSNA in both thermal conditions. However, during the heat stress trial, increases in MSNA at LBNP levels higher than -9 mmHg were greater compared with during the same LBNP levels in normothermia (all P < 0.05). These data suggest that the increase in MSNA to orthostatic stress is not attenuated but rather accentuated in heat-stressed humans.     

The effect of acute rise in intracranial pressure on the sympathetic cardiac, vertebral and phrenic nerve activities.

A transient rise of intracranial pressure in cats under chloralose-urethane anaesthesia increased the activity of the sympathetic vertebral nerve, cardiac nerve and in the first phase phrenic nerve. If the vagus nerves were intact this rise in sympathetic activity was associated with bradycardia. These effects developed with a delay, as a rule after abatement of the transient intracranial pressure rise. The authors suggest that Cushing's reaction is caused by medullary ischaemia and development of local metabolic acidosis activating simultaneously the sympathetic and parasympathetic neurons in the medulla oblongata.     

Transfer function analysis between arterial pressure and renal sympathetic nerve activity at cardiac pacing frequencies in the rat.

This study examined the possible influence of changes in heart rate (HR) on the gain of the transfer function relating renal sympathetic nerve activity (RSNA) to arterial pressure (AP) at HR frequency in rats. In seven urethane-anesthetized rats, AP and RSNA were recorded under baseline conditions (spontaneous HR = 338 +/- 6 beats/min, i.e., 5.6 +/- 0.1 Hz) and during 70-s periods of cardiac pacing at 6-9 Hz applied in random order. Cardiac pacing slightly increased mean AP (0.8 +/- 0.2 mmHg/Hz) and decreased pulse pressure (-3.6 +/- 0.3 mmHg/Hz) while leaving the mean level of RSNA essentially unaltered (P = 0.680, repeated-measures ANOVA). The gain of the transfer function from AP to RSNA measured at HR frequency was always associated with a strong, significant coherence and was stable between 6 and 9 Hz (P = 0.185). The transfer function gain measured under baseline conditions [2.44 +/- 0.28 normalized units (NU)/mmHg] did not differ from that measured during cardiac pacing (2.46 +/- 0.27 NU/mmHg). On the contrary, phase decreased linearly as a function of HR, which indicated the presence of a fixed time delay (97 +/- 6 ms) between AP and RSNA. In conclusion, the dynamic properties of arterial baroreflex pathways do not affect the gain of the transfer function between AP and RSNA measured at HR frequency in the upper part of the physiological range of HR variations in the rat.     

Neonatal maternal separation enhances phrenic responses to hypoxia and carotid sinus nerve stimulation in the adult anesthetized rat.

In awake animals, our laboratory recently showed that the hypoxic ventilatory response of adult male (but not female) rats previously subjected to neonatal maternal separation (NMS) is 25% greater than controls (Genest SE, Gulemetova R, Laforest S, Drolet G, and Kinkead R. J Physiol 554: 543-557, 2004). To begin mechanistic investigations of the effects of this neonatal stress on respiratory control development, we tested the hypothesis that, in male rats, NMS enhances central integration of carotid body chemoafferent signals. Experiments were performed on two groups of adult male rats. Pups subjected to NMS were placed in a temperature-controlled incubator 3 h/day from postnatal day 3 to postnatal day 12. Control pups were undisturbed. At adulthood (8-10 wk), rats were anesthetized (urethane; 1.6 g/kg), paralyzed, and ventilated with a hyperoxic gas mixture [inspired O2 fraction (Fi(O2)) = 0.5], and phrenic nerve activity was recorded. The first series of experiments aimed to demonstrate that NMS-related enhancement of the inspiratory motor output (phrenic) response to hypoxia occurs in anesthetized animals also. In this series, rats were exposed to moderate, followed by severe, isocapnic hypoxia (Fi(O2) = 0.12 and 0.08, respectively, 5 min each). NMS enhanced both the frequency and amplitude components of the phrenic response to hypoxia relative to controls, thereby validating the use of this approach. In a second series of experiments, NMS increased the amplitude (but not the frequency) response to unilateral carotid sinus nerve stimulation (stimulation frequency range: 0.5-33 Hz). We conclude that enhancement of central integration of carotid body afferent signal contributes to the larger hypoxic ventilatory response observed in NMS rats.     

Time-frequency coherence analysis of phrenic and hypoglossal activity in the decerebrate rat during eupnea, hyperpnea, and gasping

Fast respiratory rhythms include medium- (MFO) and high-frequency oscillations (HFO), which are much faster than the fundamental breathing rhythm. According to previous studies, HFO is characterized by high coherence (Coh) in phrenic (Ph) nerve activity, thereby providing a means of distinguishing between these two types of oscillations. Changes in Coh between the Ph and hypoglossal (XII) nerves during the transition from normal eupnic breathing to gasping have not been characterized. Experiments were performed on nine unanesthetized, chemo- and barodenervated, decerebrate adult rats, in which sustained asphyxia elicited hyperpnea and gasping. A gated time-frequency Coh analysis was developed and applied to whole Ph and medial XII nerve recordings. The results showed dynamic Ph-Ph Coh during eupnea, including MFO and HFO. XII-XII Coh during eupnea was broadband and included four distinct peaks, with low-frequency Coh dominating the epochs preceding the onset of Ph activity. During gasping, only MFO-peaks were present in Ph-Ph Coh. Bilateral XII activity showed a significant reduction in Coh and a shift toward lower frequencies during gasping. In contrast, contralateral Ph-XII Coh progressively increased during state changes from eupnea to gasping, a tendency mirrored in the startup part of the Ph activity. These data suggest significant hypoxia/hypercapnia-induced alterations in synchronization between respiratory outputs during the transition from eupnea to gasping, reflecting a reconfiguration of the respiratory network and/or alterations in the circuitry associated with the motor pools, including dynamic coupling between outputs.     

Choline kinase and ethanolamine kinase activity in the cytosol of nerve endings from rat forebrain.

Both choline kinase and ethanolamine kinase are present in the cytosol of nerve endings prepared from rat brain are the products of their action, phosphocholine (84 nmol/g fresh wt. of brain) and phosphoethanolamine (190 nmol/g fresh wt. of brain). In contrast with the enzymes from the cytosol of whole brain, both are as equally active at pH 7.5 as 9.0. Determination of kinase activity in membrane-containing tissue samples at pH9 gives low values because of the activity of alkaline phosphatase. Choline kinase, but not ethanolamine kinase, requires Mg2+ in excess of that required for the formation of the MgATP complex and is inhibited by an excess of free ATP. The Km for choline is 2.6mM and for ethanolamine is 2.2mM. The differing requirements for ATP and Mg2+ and the inhibition of choline kinase, but not ethanolamine kinase, by hemicholinium-3 suggest either the presence of two separate enzymes or two different active sites on the same enzyme.     

Expression of nerve growth factor receptor immunoreactivity in the rat choroid plexus.

The presence and localization of nerve growth factor receptors (NGFr) in the choroid plexus of the adult rat has been investigated immunohistochemically using an anti-rat NGFr monoclonal antibody (192-IgG). A moderate to strong immunoreaction was observed in the epithelial cells of the choroid plexus, whereas the choroidal blood vessels and connective tissue remained unlabelled. Moreover, no sex-differences were encountered in the NGFr immunoreaction intensity and Bouin fixative was more effective than 10% formaldehyde evidenciating the NGFr immunostain. Occasionally, ependymal cells displaying NGFr immunoreactivity were observed. Present data demonstrate that the choroid plexus of the rat contain NGFr, probably low-affinity NGFr, and suggest an involvement of NGF in the regulation of cerebrospinal fluid secretion, but the importance of these findings, if any, must be investigated in future studies.