Strain differences in pH-sensitive K+ channel-expressing cells in chemosensory and nonchemosensory brain stem nuclei.

The ventilatory CO2 chemoreflex is inherently low in inbred Brown Norway (BN) rats compared with other strains, including inbred Dahl salt-sensitive (SS) rats. Since the brain stem expression of various pH-sensitive ion channels may be determinants of the CO2 chemoreflex, we tested the hypothesis that there would be fewer pH-sensitive K(+) channel-expressing cells in BN relative to SS rats within brain stem sites associated with respiratory chemoreception, such as the nucleus tractus solitarius (NTS), but not within the pre-Bötzinger complex region, nucleus ambiguus or the hypoglossal motor nucleus. Medullary sections (25 µm) from adult male and female BN and SS rats were stained with primary antibodies targeting TASK-1, Kv1.4, or Kir2.3 K(+) channels, and the total (Nissl-stained) and K(+) channel immunoreactive (-ir) cells counted. For both male and female rats, the numbers of K(+) channel-ir cells within the NTS were reduced in the BN compared with SS rats (P < 0.05), despite equal numbers of total NTS cells. In contrast, we found few differences in the numbers of K(+) channel-ir cells among the strains within the nucleus ambiguus, hypoglossal motor nucleus, or pre-Bötzinger complex regions in both male and female rats. However, there were no predicted functional mutations in each of the K(+) channels studied comparing genomic sequences among these strains. Thus we conclude that the relatively selective reductions in pH-sensitive K(+) channel-expressing cells in the NTS of male and female BN rats may contribute to their severely blunted ventilatory CO2 chemoreflex.

Contributions of the Kölliker-Fuse nucleus to coordination of breathing and swallowing.

Herein we compare the effects of perturbations in the Kölliker-Fuse nucleus (KFN) and the lateral (LPBN) and medial (MPBN) parabrachial nuclei on the coordination of breathing and swallowing. Cannula was chronically implanted in goats through which ibotenic acid (IA) was injected while awake. Swallows in late expiration (E) always reset while swallows in early inspiration (I) never reset the respiratory rhythm. Before cannula implantation, all other E and I swallows did not reset the respiratory rhythm, and had small effects on E and I duration and tidal volume (VT). However, after cannula implantation in the MPBN and KFN, E and I swallows reset the respiratory rhythm and increased the effects on I and E duration and VT. Subsequent injection of IA into the KFN eliminated the respiratory phase resetting of swallows but exacerbated the effects on I and E duration and VT. We conclude that the KFN and to a lesser extent the MPBN contribute to coordination of breathing and swallowing.

Postnatal ventilatory response to CO2 in awake piglets.

Abnormal ventilatory responses to increased levels of inspired CO2 during postnatal development may pose a risk for Sudden Infant Death Syndrome, primarily during periods of vulnerability. The purpose of this study was to test the hypothesis that in awake piglets the ventilatory response to hypercapnia would be attenuated between 10 and 15 days of age relative to younger and older ages. To test this hypothesis, we measured the ventilatory response to 5% inspired CO2 in piglets from postnatal (PN) days 1 through PN28. Piglets were divided into groups and exposed to 5% CO2 daily, every 3rd day or on and after PN20-21 only to avoid any plasticity that may result from repeated exposure to CO2. Room air ventilation normalized to body weight (V˙(E), ml/min/kg) declined with postnatal age in piglets from all groups. The ventilatory response to 5% inspired CO2 (expressed as % change from control) was present at birth, and we did not find an age-dependent change from PN1 to PN28 (p > 0.1). In addition, we did not find that repeated exposure (daily or every 3rd day) to 5% inspired CO2 altered the ventilatory response during this period of development. We conclude that the previously documented apparent critical period of development in piglets between 10 and 15 days of age is not associated with attenuation of the ventilatory response to 5% inspired CO2.

The effects of lesions in the dorsolateral pons on the coordination of swallowing and breathing in awake goats.

The purpose of this retrospective study was to gain insight into the contribution of the dorsolateral pons to the coordination of swallowing and breathing in awake goats. In 4 goats, cannulas were chronically implanted bilaterally through the lateral (LPBN) and medial (MPBN) parabrachial nuclei just dorsal to the Kölliker-Fuse nucleus (KFN). After >2weeks recovery from this surgery, the goats were studied for 5½h on a control day, and on separate days after receiving 1 and 10µl injections of ibotenic acid (IA) separated by 1week. The frequency of swallows did not change during the control and 1µl IA studies, but after injection of 10µl IA, there was a transient 65% increase in frequency of swallows (P<0.05). Under control conditions swallows occurred throughout the respiratory cycle, where late-E swallows accounted for 67.6% of swallows. The distribution of swallow occurrence throughout the respiratory cycle was unaffected by IA injections. Consistent with the concept that swallowing is dominant over breathing, we found that swallows increased inspiratory (T(I)) and expiratory (T(E)) time and decreased tidal volume (V(T)) of the breath of the swallow (n) and/or the subsequent (n+1) breath. Injections of 10µl IA attenuated the normal increases in T(I) and T(E) and further attenuated V(T) of the n breath. Additionally, E and I swallows reset respiratory rhythm, but injection of 1 or 10µl IA progressively attenuated this resetting, suggesting a decreased dominance over respiratory motor output with increasing IA injections. Post mortem histological analysis revealed about 50% fewer (P<0.05) neurons remained in the KFN, LPBN, and MPBN in lesioned compared to control goats. We conclude that dorsolateral pontine nuclei have a modulatory role in a hypothesized holarchical neural network regulating swallowing and breathing particularly contributing to the normal dominance of swallowing over breathing in both rhythm and motor pattern generation.

Anatomic changes in multiple brainstem nuclei after incremental, near-complete neurotoxic destruction of the pre-Bötzinger Complex in adult goats.

Abrupt, bilateral destruction of the pre-Bötzinger Complex (preBötC) leads to terminal apnea in unanesthetized goats and rats. In contrast, respiratory rhythm and pattern and arterial blood gases in goats during wakefulness and sleep are normal after incremental (over a month) destruction of > 90% of the preBötC. Here, we tested the hypothesis that the difference in effects between abrupt and incremental destruction of the preBötC are a result of time-dependent plasticity, which manifests as anatomic changes at sites within the respiratory network. Accordingly, we report data from histological analyses comparing the brainstems of control goats, and goats that had undergone bilateral, incremental, ibotenic acid (IA)-induced preBötC lesioning. A major focus was on the parafacial respiratory group/retrotrapezoid nucleus (pFRG/RTN) and the pontine respiratory group (PRG), which are sites thought to contribute to respiratory rhythmogenesis. We also studied the facial (FN), rostral nucleus ambiguus (NA), medullary raphé (MRN), hypoglossal (HN), and the dorsal motor vagal (DMV) nuclei. Neuronal counts, count region area (mm²), and neuronal densities were calculated using computer-assisted analyses and/or manual microscopy to compare control and preBötC-lesioned animals. We found that within the ventral and lateral medulla 2mm rostral to the caudal pole of the FN (presumed pFRG/RTN), there were 25% and 65% more (P < 0.001) neurons, respectively, in preBötC-lesioned compared to control goats. Lesioned goats also showed 14% and 13% more (P < 0.001) neurons in the HN and medial parabrachialis nucleus, but 46%, 28%, 7%, and 17% fewer (P < 0.001) neurons in the FN, NA, DMV, and Kölliker-Fuse nuclei, respectively. In the remaining sites analyzed, there were no differences between groups. We conclude that anatomic changes at multiple sites within the respiratory network may contribute to the time-dependent plasticity in breathing following incremental and near-complete destruction of the preBötC.

Site-specific effects on respiratory rhythm and pattern of ibotenic acid injections in the pontine respiratory group of goats.

To probe further the contributions of the rostral pons to eupneic respiratory rhythm and pattern, we tested the hypothesis that ibotenic acid (IA) injections in the pontine respiratory group (PRG) would disrupt eupneic respiratory rhythm and pattern in a site- and state-specific manner. In 15 goats, cannulas were bilaterally implanted into the rostral pontine tegmental nuclei (RPTN; n = 3), the lateral (LPBN; n = 4) or medial parabrachial nuclei (MPBN; n = 4), or the Kölliker-Fuse nucleus (KFN; n = 4). After recovery from surgery, 1- and 10-microl injections (1 wk apart) of IA were made bilaterally through the implanted cannulas during the day. Over the first 5 h after the injections, there were site-specific ventilatory effects, with increased (P < 0.05) breathing frequency in RPTN-injected goats, increased (P < 0.05) pulmonary ventilation (Vi) in LPBN-injected goats, no effect (P < 0.05) in MPBN-injected goats, and a biphasic Vi response (P < 0.05) in KFN-injected goats. This biphasic response consisted of a hyperpnea for 30 min, followed by a prolonged hypopnea and hypoventilation with marked apneas, apneusis-like breathing patterns, and/or shifts in the temporal relationships between inspiratory flow and diaphragm activity. In the awake state, 10-15 h after the 1-microl injections, the number of apneas was greater (P < 0.05) than during other studies at night. However, there were no incidences of terminal apneas. Breathing rhythm and pattern were normal 22 h after the injections. Subsequent histological analysis revealed that for goats with cannulas implanted into the KFN, there were nearly 50% fewer neurons (P < 0.05) in all three PRG subnuclei than in control goats. We conclude that in awake goats, 1) IA injections into the PRG have site-specific effects on breathing, and 2) the KFN contributes to eupneic respiratory pattern generation.

A role for the Kolliker-Fuse nucleus in cholinergic modulation of breathing at night during wakefulness and NREM sleep.

For many years, acetylcholine has been known to contribute to the control of breathing and sleep. To probe further the contributions of cholinergic rostral pontine systems in control of breathing, we designed this study to test the hypothesis that microdialysis (MD) of the muscarinic receptor antagonist atropine into the pontine respiratory group (PRG) would decrease breathing more in animals while awake than while in NREM sleep. In 16 goats, cannulas were bilaterally implanted into rostral pontine tegmental nuclei (n = 3), the lateral (n = 3) or medial (n = 4) parabrachial nuclei, or the Kölliker-Fuse nucleus (KFN; n = 6). After >2 wk of recovery from surgery, the goats were studied during a 45-min period of MD with mock cerebrospinal fluid (mCSF), followed by at least 30 min of recovery and a second 45-min period of MD with atropine. Unilateral and bilateral MD studies were completed during the day and at night. MD of atropine into the KFN at night decreased pulmonary ventilation and breathing frequency and increased inspiratory and expiratory time by 12-14% during both wakefulness and NREM sleep. However, during daytime studies, MD of atropine into the KFN had no effect on these variables. Unilateral and bilateral nighttime MD of atropine into the KFN increased levels of NREM sleep by 63 and 365%, respectively. MD during the day or at night into the other three pontine sites had minimal effects on any variable studied. Finally, compared with MD of mCSF, bilateral MD of atropine decreased levels of acetylcholine and choline in the effluent dialysis fluid. Our data support the concept that the KFN is a significant contributor to cholinergically modulated control of breathing and sleep.

The pontine respiratory group, particularly the Kölliker-Fuse nucleus, mediates phases of the hypoxic ventilatory response in unanesthetized goats.

The objective of the present study was to test the hypothesis that, in the in vivo awake goat model, perturbation/lesion in the pontine respiratory group (PRG) would decrease the sensitivity to hypercapnia and hypoxia. The study reported herein was part of two larger studies in which cholinergic modulation in the PRG was attenuated by microdialysis of atropine and subsequently ibotenic acid injections neurotoxically lesioned the PRG. In 14 goats, cannula were bilaterally implanted into either the lateral (n=4) or medial (n=4) parabrachial nuclei or the Kölliker-Fuse nucleus (KFN, n=6). Before and after cannula implantation, microdialysis of atropine, and injection of ibotenic acid, hypercapnic and hypoxic ventilatory sensitivities were assessed. Hypercapnic sensitivity was assessed by three 5-min periods at 3, 5, and 7% inspired CO2. In all groups of goats, CO2 sensitivity was unaffected (P>0.05) by any PRG perturbations/lesions. Hypoxic sensitivity was assessed with a 30-min period at 10.8% inspired O2. The response to hypoxia was typically triphasic, with a phase 1 increase in pulmonary ventilation, a phase 2 roll-off, and a phase 3 prolonged increase associated with shivering and increased metabolic rate and body temperature. In all groups of goats, the phase 1 of the hypoxic ventilatory responses was unaffected by any PRG perturbations/lesions, and there were no consistent effects on the phase 2 responses. However, in the KFN group of goats, the phase 3 ventilatory, shivering, metabolic rate, and temperature responses were markedly attenuated after the atropine dialysis studies, and the attenuation persisted after the ibotenic acid studies. These findings support an integrative or modulatory role for the KFN in the phase 3 responses to hypoxia.

Contributions of central and peripheral chemoreceptors to the ventilatory response to CO2/H+.

The major objective of this review is to evaluate existing information and reach conclusions regarding whether there is interaction between P(CO(2))/H(+) stimulation of carotid (peripheral) and intracranial (central) chemoreceptors. Interaction is defined as a ventilatory response to simultaneous changes in the degree of Pco2/H(+) stimulation of both chemoreceptors that is greater (hyperadditive) or less (hypoadditive) than the sum of the responses when stimulation of each set of chemoreceptors is individually altered. Simple summation of the simultaneous changes in stimuli results in no interaction (i.e., additive interaction). Knowledge of the nature of central/peripheral interaction is crucial for determining the physiological significance of newer models of ventilatory control based on recent neuroanatomic observations of the circuitry of key elements of the ventilatory control system. In this review, we will propose that these two sets of receptors are not functionally separate but rather that they are dependent on one another such that the sensitivity of the medullary chemoreceptors is critically determined by input from the peripheral chemoreceptors and possibly other breathing-related reflex afferents as well. The short format of this minireview demands that we be somewhat selective in developing our ideas. We will briefly discuss the limitations of experiments used to study CO(2)/H(+) sensitivity and interaction to date, traditional views of the relative contributions of peripheral and central chemoreceptors to CO(2)/H(+) sensitivity, the evidence for and against different types of interaction, and the effect of tonic carotid chemoreceptor afferent activity on central control mechanisms.

Differences between three inbred rat strains in number of K+ channel-immunoreactive neurons in the medullary raphé nucleus.

Ventilatory sensitivity to hypercapnia is greater in Dahl salt-sensitive (SS) rats than in Fawn Hooded hypertensive (FHH) and Brown Norway (BN) inbred rats. Since pH-sensitive potassium ion (K(+)) channels are postulated to contribute to the sensing and signaling of changes in CO(2)-H(+) in chemosensitive neurons, we tested the hypothesis that there are more pH-sensitive K(+) channel-immunoreactive (ir) neurons within the medullary raphé nuclei of the highly chemosensitive SS rats than in the other two strains. Medullary tissues from male and female BN, FHH, and SS rats were stained with cresyl violet or with antibodies targeting TASK-1, K(v)1.4, and Kir2.3 channels. K(+) channel-ir neurons were quantified and compared with the total neurons in the region. The total number of neurons in the medullary raphé 1) was greater in male FHH than the other male rats, 2) did not differ among the female rats, and 3) did not differ between sexes. The average number of K(+) channel-ir neurons per section was 30-60 neurons higher in the male SS than in the other rat strains. In contrast, for the females, the number of K(+) channel-ir neurons was greatest in the BN. We also found significant differences in the number of K(+) channel-ir neurons between sexes in SS (males > females) and BN (females > males) rats, but not the FHH strain. Our findings support the hypothesis for males but not for females, suggesting that both genetic background and sex are determinants of K(+) channel immunoreactivity of medullary raphé neurons, and that the expression of pH-sensitive K(+) channels in the medullary raphé does not correlate with the ventilatory sensitivity to hypercapnia.

Micro-opioid receptor agonist injections into the presumed pre-Botzinger complex and the surrounding region of awake goats do not alter eupneic breathing.

Opioids are clinically important in the alleviation of pain. An undesirable side effect of opioids is depression of breathing. Data from isolated preparations suggest this effect is due to attenuation of discharge activity of neurons in the pre-Bötzinger complex (preBötzC), a medullary area with respiratory rhythmogenic properties. The purpose of this study was to examine how [d-Ala(2),N-Me-Phe(4),Gly(5)-ol]-enkephalin (DAMGO), a mu-opioid receptor agonist, affected breathing after injection into the presumed preBötzC of the adult awake goat. We hypothesized that DAMGO would cause breathing to decrease and become irregular when injected into the presumed preBötzC and the surrounding region of the conscious animal. We further hypothesized that ventilatory sensitivity to CO(2) and hypoxia would be blunted after the injection of DAMGO. Microtubules were bilaterally implanted into the presumed preBötzC of 10 adult female goats. After recovery from the surgery, DAMGO (0.5-10 mul, 1 nM-10 muM) was injected into the presumed preBötzC during the awake state. DAMGO had no effect on pulmonary ventilation [inspiratory minute ventilation (Vi)], respiratory rhythm and pattern, the activation pattern of inspiratory and expiratory muscles, or arterial blood gases during eupneic breathing conditions (P > 0.10). However, DAMGO attenuated (P < 0.05) the evoked increase in breathing frequency when inspired CO(2) was increased, and DAMGO attenuated the Vi response to reduction of inspired O(2) to 10.8% (P < 0.05). We conclude that our data do not provide support for the concept that in awake mammals opioid depression of breathing is due to a directed action of opioids on preBötzC neurons.

Normal breathing pattern and arterial blood gases in awake and sleeping goats after near total destruction of the presumed pre-Botzinger complex and the surrounding region.

Abrupt neurotoxic destruction of >70% of the pre-Bötzinger complex (preBötzC) in awake goats results in respiratory and cardiac failure (Wenninger JM, Pan LG, Klum L, Leekley T, Bastastic J, Hodges MR, Feroah TR, Davis S, Forster HV. J Appl Physiol 97: 1629-1636, 2004). However, in reduced preparations, rhythmic respiratory activity has been found in other areas of the brain stem (Huang Q, St. John WM. J Appl Physiol 64: 1405-1411, 1988; Janczewski WA, Feldman JL. J Physiol 570: 407-420, 2006; Lieske SP, Thoby-Brisson M, Telgkamo P, Ramierz JM. Nature Neurosci 3: 600-607, 2000; St. John WM, Bledsoe TA. J Appl Physiol 59: 684-690, 1985); thus we hypothesized that, when the preBötzC is destroyed incrementally over weeks, time-dependent plasticity within the respiratory network will result in a respiratory rhythm capable of maintaining normal blood gases. Microtubules were bilaterally implanted into the presumed preBötzC of seven goats. After recovery from surgery, studies were completed to establish baseline values for respiratory parameters. At weekly intervals, increasing volumes (in order 0.5, 1, 5, and 10 microl) of ibotenic acid (IA; 50 mM) were then injected into the preBötzC. All IA injections resulted in an acute tachypnea and dysrhythmia featuring augmented breaths, apneas, and increased breath-to-breath variation in breathing. In studies at night, apneas were nearly all central and occurred in the awake state. Breath-to-breath variation in breathing was greater (P < 0.05) during wakefulness than during non-rapid eye movement sleep. However, one week after the final IA injection, the breathing pattern, breath-to-breath variation, and arterial blood gases and pH were unchanged from baseline, but there was a 20% decrease in respiratory frequency (f) and CO(2) sensitivity (P < 0.05), as well as a 40% decrease in the ventilatory response to hypoxia (P < 0.001). In subsequent histological analysis of the presumed preBötzC region of lesioned goats, it was determined that there was a 90 and 92% reduction from control goats in total and neurokinin-1 receptor neurons, respectively. Therefore, it was concluded that 1) the dysrhythmic effects on breathing are state dependent; and 2) after incremental, near total destruction of the presumed preBötzC region, time-dependent plasticity within the respiratory network provides a rhythm capable of sustaining normal arterial blood gases.

Focal acidosis in the pre-Botzinger complex area of awake goats induces a mild tachypnea.

There are widespread chemosensitive areas in the brain with varying effects on breathing. In the awake goat, microdialyzing (MD) 50% CO(2) at multiple sites within the medullary raphe increases pulmonary ventilation (Vi), blood pressure, heart rate, and metabolic rate (Vo(2)) (11), while MD in the rostral and caudal cerebellar fastigial nucleus has a stimulating and depressant effect, respectively, on these variables (17). In the anesthetized cat, the pre-Bötzinger complex (preBötzC), a hypothesized respiratory rhythm generator, increases phrenic nerve activity after an acetazolamide-induced acidosis (31, 32). To gain insight into the effects of focal acidosis (FA) within the preBötzC during physiological conditions, we tested the hypothesis that FA in the preBötzC during wakefulness would stimulate breathing, by increasing respiratory frequency (f). Microtubules were bilaterally implanted into the preBötzC of 10 goats. Unilateral MD of mock cerebral spinal fluid equilibrated with 6.4% CO(2) did not affect Vi, tidal volume (Vt), or f. Unilateral MD of 25 and 50% CO(2) significantly increased Vi and f by 10% (P < 0.05, n = 10, 17 trials), but Vt was unaffected. Bilateral MD of 6.4, 25, or 50% CO(2) did not significantly affect Vi, Vt, or f (P > 0.05, n = 6, 6 trials). MD of 80% CO(2) caused a 180% increase in f and severe disruptions in airflow (n = 2). MD of any level of CO(2) did not result in any significant changes in mean arterial blood pressure, heart rate, or Vo(2). Thus the data suggest that the preBötzC area is chemosensitive, but the responses to FA at this site are unique compared with other chemosensitive sites.

The cerebellar fastigial nucleus contributes to CO2-H+ ventilatory sensitivity in awake goats.

The purpose of this study was to test the hypothesis that an intact cerebellar fastigial nucleus (CFN) is an important determinant of CO(2)-H(+) sensitivity during wakefulness. Bilateral, stainless steel microtubules were implanted into the CFN (N=9) for injection (0.5-10 microl) of the neurotoxin ibotenic acid. Two or more weeks after implantation of the microtubules, eupneic breathing and CO(2)-H(+) sensitivity did not differ significantly (P>0.10) from pre-implantation conditions. Injection of ibotenic acid (50 mM) did not significantly alter eupneic Pa(CO2) (P>0.10). The coefficient of variation of eupneic Pa(CO2) was 4.0+/-0.6 and 3.7+/-0.4% over the 2 weeks before and after the lesion, respectively. CO(2)-H(+) sensitivity expressed as inspired ventilation/Pa(CO2) decreased from 2.15+/-0.17 pre-lesion to 1.58+/-0.26 l/(min mmHg) 3-6 days post-lesion (P<0.02, -27%). There was no significant (P>0.10) recovery of sensitivity between 7 and 10 days post-lesion. The lesion also increased (P<0.05) the day-to-day variability of this index by nearly 100%. When CO(2) sensitivity was expressed as elevated inspired CO(2)/room air V (I), values at 7%, but not 3 and 5% inspired CO(2), were reduced and more variable (P<0.05) after the ibotenic acid injections. We conclude that during wakefulness, the CFN contributes relatively more to overall ventilatory drive at high relative to low levels of hypercapnia.

Lesions in the cerebellar fastigial nucleus have a small effect on the hyperpnea needed to meet the gas exchange requirements of submaximal exercise.

The purpose of this study was to test the hypothesis that an intact cerebellar fastigial nucleus (CFN) is necessary for the hyperpnea to meet the gas exchange needs of submaximal exercise. Bilateral stainless steel microtubules were implanted in the cerebellum inside (n = 12) or outside (n = 2) the CFN for injection (0.5 to 10 microl) of the neurotoxin ibotenic acid. All goats had difficulty maintaining normal posture and walking for up to 1 mo after the implantation of the microtubules and again for hours or days after the neurotoxin was injected. Postmortem histology indicated there were 55% fewer living neurons (P < 0.001, n = 9, 3,720 +/- 553 vs. 1,670 +/- 192) in the CFN of the experimental goats compared with a control group of goats. As is typical for goats before implantation of the microtubules, the decrease in arterial Pco(2) from rest during mild and moderate treadmill exercise was 2.0 +/- 0.39 and 3.5 +/- 0.45 Torr, respectively. Implantation of the microtubules did not significantly change this exercise hyperventilation. However, neurotoxic lesioning with 10 mul ibotenic acid significantly (P < 0.05) attenuated the decrease in arterial Pco(2) by 1.3 and 2.8 Torr at the first and second workload, respectively. The modest attenuation of the exercise hypocapnia at both workloads in CFN-lesioned goats suggests that the CFN is part of the control system that enables the ventilatory response to meet the gas exchange requirements of submaximal exercise.

Postnatal developmental changes in CO2 sensitivity in rats.

Ventilatory sensitivity to CO(2) in awake adult Brown Norway (BN) rats is 50-75% lower than in adult Sprague-Dawley (SD) and salt-sensitive Dahl S (SS) rats. The purpose of the present study was to test the hypothesis that this difference would be apparent during the development of CO(2) sensitivity. Four litters of each strain were divided into four groups such that rats were exposed to 7% inspired CO(2) for 5 min in a plethysmograph every third day from postnatal day (P) 0 to P21 and again on P29 and P30. From P0 to P14, CO(2) exposure increased pulmonary ventilation (Ve) by 25-50% in the BN and SD strains and between 25 to over 200% in the SS strain. In all strains beginning around P15, the response to CO(2) increased progressively reaching a peak at P19-21 when Ve during hypercapnia was 175-225% above eucapnia. There were minimal changes in CO(2) sensitivity between P21 and P30, and at both ages there were minimal between-strain differences. At P30, the response to CO(2) in the SS and SD strains was near the adult response, but the response in the BN rats was 100% greater at P30 than in adults. We conclude that 1) CO(2)-sensing mechanisms, and/or mechanisms downstream from the chemoreceptors, change dramatically at the age in rats when other physiological systems are also maturing ( approximately P15), and 2) there is a high degree of age-dependent plasticity in CO(2) sensitivity in rats, which differs between strains.

CO2/H+ chemoreceptors in the cerebellar fastigial nucleus do not uniformly affect breathing of awake goats.

Our objective in this study was to test the hypothesis that focal acidosis (FA) in the cerebellar fastigial nucleus (CFN) of awake goats arising from global brain acidosis induced by increasing inspired CO2 will increase breathing. FA was created by reverse microdialysis of mock cerebral spinal fluid, equilibrated with 6.4, 25, 50, or 80% CO2 through chronically implanted microtubules (cannula). Dialysis with 6.4% CO2 had no significant effects on any physiological parameters. However, microdialysis at higher levels of CO2 increased pulmonary ventilation (V(I)) in one group of studies and decreased V(I) in a second group and the difference between the groups was significant (t = 9.16, P < 0.001). In one group of studies (n = 8), FA with 50 and 80% CO2 significantly increased (P < 0.05) Vi by 16 and 12%, respectively, and significantly increased (P < 0.05) heart rate by 13 and 9%, respectively. In contrast, in another group of studies (n = 6), FA with 25 and 50% CO2 significantly decreased (P < 0.05) Vi by 7 and 10%, respectively. In this group oxygen consumption was decreased during dialysis with 80% CO2. On the basis of histology, we estimate that the increased and decreased responses were associated with FA primarily in the rCFN and cCFN, respectively. We conclude that there are CO2/H+-sensitive neurons in the CFN that do not uniformly affect breathing. In addition, the significant changes in heart rate and oxygen consumption during FA indicate that the CFN can also influence non-respiratory-related control systems.

Genetic determinants on rat chromosome 6 modulate variation in the hypercapnic ventilatory response using consomic strains.

To understand the genetic basis of pathways involved in the control of breathing, a large scale, high-throughput study using chromosomal substitution strains of rats is underway. Eight new consomic rat stains (SS-2(BN), SS-4(BN), SS-6(BN), SS-7(BN), SS-8(BN), SS-11(BN), SS-12(BN), SS-14(BN), SS-Y(BN)), containing one homozygous BN/NHsdMcwi (BN) chromosome on a background of SS/JrHsdMcwi (SS), were created by PhysGen (http://pga.mcw.edu) Program for Genomic Applications. Male and female rats were studied using standard plethysmography under control conditions and during acute hypoxia (inspired oxygen fraction = 0.12) and hypercapnia (inspired CO(2) fraction = 0.07). The rats were also studied during treadmill exercise. Both male and female BN rats had a significantly lower ventilatory response during 7% CO(2) compared with SS rats of the same gender. SS-6(BN) female rats had a significantly reduced ventilatory response, similar to BN rats due primarily to a reduced tidal volume. Male SS-6(BN) rats had a significantly reduced tidal volume response to hypercapnia but a slightly increased frequency response during hypercapnia. Gene(s) on the Y chromosome may play a role in this increased frequency response in the male rats because the SS-Y(BN) hypercapnic ventilatory response involves a significantly increased frequency response. Several chromosomal substitutions slightly altered the ventilatory responses to hypoxia and exercise. However, genes on chromosomes 6 and Y of those studied are of primary importance in aspects of ventilatory control currently studied.

Carotid body denervation alters ventilatory responses to ibotenic acid injections or focal acidosis in the medullary raphe.

Our aim was to determine the effects of carotid body denervation (CBD) on the ventilatory responses to focal acidosis and ibotenic acid (IA) injections into the medullary raphe area of awake, adult goats. Multiple microtubules were chronically implanted into the midline raphe area nuclei either before or after CBD. For up to 15 days after bilateral CBD, arterial PCO2 (PaCO2) (13.3 +/- 1.9 Torr) was increased (P < 0.001), and CO2 sensitivity (-53.0 +/- 6.4%) was decreased (P <0.001). Thereafter, resting PaCO2 and CO2 sensitivity returned (P <0.01) toward control, but PaCO2 remained elevated (4.8 +/- 1.9 Torr) and CO2 sensitivity reduced (-24.7 +/- 6.0%) > or =40 days after CBD. Focal acidosis (FA) at multiple medullary raphe area sites 23-44 days post-CBD with 50 or 80% CO(2) increased inspiratory flow (Vi), tidal volume (Vt), metabolic rate (VO2), and heart rate (HR) (P <0.05). The effects of FA with 50% CO2 after CBD did not differ from intact goats. However, CBD attenuated (P <0.05) the increase in Vi, Vt, and HR with 80% CO2, but it had no effect on the increase in VO2. Rostral but not caudal raphe area IA injections increased Vi, BP, and HR (P < 0.05), and these responses were accentuated (P <0.001) after CBD. CO2 sensitivity was attenuated (-20%; P <0.05) <7 days after IA injection, but thereafter it returned to prelesion values in CBD goats. We conclude the following: 1) the attenuated response to FA after CBD provides further evidence that the carotid bodies provide a tonic facilitory input into respiratory control centers, 2) the plasticity after CBD is not due to increased raphe chemoreceptor sensitivity, and 3) the "error-sensing" function of the carotid body blunts the effect of strong stimulation of the raphe.