. Transient attenuation of CO 2 sensitivity after neurotoxic lesions in the medullary raphe area of awake goats. J Appl Physiol 97: 2236 -2247, 2004. First published August 20, 2004; doi:10.1152/japplphysiol.00584.2004The major objective of this study was to gain insight into whether under physiological conditions medullary raphe area neurons influence breathing through CO 2/H ϩ chemoreceptors and/or through a postulated, nonchemoreceptor modulatory influence. Microtubules were chronically implanted into the raphe of adult goats (n ϭ 13), and breathing at rest (awake and asleep), breathing during exercise, as well as CO 2 sensitivity were assessed repeatedly before and after sequential injections of the neurotoxins saporin conjugated to substance P [SP-SAP; neurokinin-1 receptor (NK1R) specific] and ibotenic acid (IA; nonspecific glutamate receptor excitotoxin). In all goats, microtubule implantation alone resulted in altered breathing periods, manifested as central or obstructive apneas, and fractionated breathing. The frequency and characteristics of the altered breathing periods were not subsequently affected by injections of the neurotoxins (P Ͼ 0.05). Three to seven days after SP-SAP or subsequent IA injection, CO 2 sensitivity was reduced (P Ͻ 0.05) by 23.8 and 26.8%, respectively, but CO 2 sensitivity returned to preinjection control values Ͼ7 days postinjection. However, there was no hypoventilation at rest (awake, non-rapid eye movement sleep, or rapid eye movement sleep) or during exercise after these injections (P Ͼ 0.05). The neurotoxin injections resulted in neuronal death greater than three times that with microtubule implantation alone and reduced (P Ͻ 0.05) both tryptophan hydroxylase-expressing (36%) and NK1R-expressing (35%) neurons at the site of injection. We conclude that both NK1R-and glutamate receptor-expressing neurons in the medullary raphe nuclei influence CO 2 sensitivity apparently through CO 2/H-expressing chemoreception, but the altered breathing periods appear unrelated to CO 2 chemoreception and thus are likely due to non-chemoreceptor-related neuromodulation of ventilatory control mechanisms. central chemoreception; control of breathing THE MEDULLARY RAPHE NUCLEI project to and supposedly modulate several central nervous system regions, including the pre-Bötzinger complex; the hypoglossal, phrenic, and spinal motoneurons; the nucleus ambiguus; the nucleus of the solitary tract; and various pontine nuclei (3,4,8,16,(37)(38)(39). The neuronal population in the raphe is heterogeneous, containing neurokinin-1 receptor-expressing (NK1R) and serotonergic neurons (28). Serotonergic neurons corelease thyrotropin-releasing hormone and substance P (SP), which are all capable of modulating ventilation (5, 7, 14, 15, 19 -21). Therefore, these raphe neurons are capable of influencing breathing through multiple neuromodulators acting at multiple levels, including the rhythm-and pattern-generating neurons and motoneurons. The primary site of raphe neuromodulation during physiological condition...

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.

Krause KL, Forster HV, Davis SE, Kiner T, Bonis JM, Pan LG, Qian B. Focal acidosis in the pre-Bötzinger complex area of awake goats induces a mild tachypnea. J Appl Physiol 106: 241-250, 2009. First published November 13, 2008 doi:10.1152/japplphysiol.90547.2008There are widespread chemosensitive areas in the brain with varying effects on breathing. In the awake goat, microdialyzing (MD) 50% CO2 at multiple sites within the medullary raphe increases pulmonary ventilation (V I), blood pressure, heart rate, and metabolic rate (V O2) (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% CO2 did not affect V I, tidal volume (VT), or f. Unilateral MD of 25 and 50% CO2 significantly increased V I and f by 10% (P Ͻ 0.05, n ϭ 10, 17 trials), but VT was unaffected. Bilateral MD of 6.4, 25, or 50% CO2 did not significantly affect V I, VT, or f (P Ͼ 0.05, n ϭ 6, 6 trials). MD of 80% CO2 caused a 180% increase in f and severe disruptions in airflow (n ϭ 2). MD of any level of CO2 did not result in any significant changes in mean arterial blood pressure, heart rate, or V O2. 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.breathing; chemosensitivity CENTRAL RESPIRATORY CO 2 /H ϩ chemoreception has been traditionally attributed to sites at or near the ventrolateral medullary surface (16,24,27). However, studies over the last 15-20 yr indicate that the ventrolateral medullary surface is not the sole CO 2 /H ϩ -sensitive area (1-3, 5, 15, 17, 21, 24, 31, 32). Electrophysiological recordings of in vitro preparations have shown that neurons in the retrotrapezoid nucleus, nucleus of the solitary tract, the medullary raphe nucleus (MRN), locus coeruleus, and the pre-Bötzinger complex (preBötzC) increase discharge frequency when the pH in the bathing solution is reduced (3,5,8,14,22,24). Studies in anesthetized preparations that either created a focal acidosis (FA) or assessed the effects of lesions on the hypercapnic ventilatory response also support the concept of widespread chemosensitivity in the medulla, pons, and cerebellum (11,18,32,33,35,36,37).While the in vivo data, under physiological conditions, suggest that chemoreceptors at multiple sites influence breathing, the observed ventilatory effects are not uniform, and the hyperpnea is small, with FA at a single si...

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.

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.