Gerald E. Bisgard

Carotid body mechanisms in acclimatization to hypoxia

Most studies oriented toward examining mechanisms increasing carotid body (CB) sensitivity to hypoxia during ventilatory acclimatization (VAH) have focussed on the role of known neuromodulators of CB function. Two general categories of the neuromodulatory agents studied most extensively could be considered: those thought to be primarily inhibitory to CB function: dopamine, norepinephrine, nitric oxide and those thought to be primarily excitatory: substance P, endothelin. There is evidence that these putative inhibitory agents are up-regulated in the first weeks of chronic hypoxia and that substance P is down-regulated. All these changes would favor a decrease in CB sensitivity to hypoxia. There are data suggesting that CB endothelin activity is up-regulated in rats subjected to chronic hypoxia, a direction suggesting increased CB sensitivity to hypoxia. Dopamine may have an excitatory as well as an inhibitory role on the CB, but there is not yet evidence to indicate that an excitatory role for DA exists in chronic hypoxia. Ion channel studies of type I CB cells suggest increased excitability after prolonged hypoxia. The role of excitatory CB nicotinic receptors and putative serotonin type 3 receptors should be examined further for their potential role in VAH. It is suggested that a balance of excitatory and inhibitory modulation is responsible for increased CB sensitivity to hypoxia during VAH.

Postnatal growth of the carotid body

The size of the carotid body (CB) is increased significantly during the postnatal period. Type I cells in the CB are the chemoreceptive element and possess many neuron-like characteristics. In contrast to previous opinions that the number of type I cells is determined before birth, we have found that type I cells continue to proliferate over a period of at least 1 month after birth in rats. The proliferation of type I cells is influenced by oxygen concentration in ambient air. Specifically, hyperoxia inhibits the type I cell proliferation, resulting in small CBs throughout life and the permanent impairment of CB chemoreception. On the other hand, hypoxia enhances the type I cell proliferation. Whether hypoxia causes long-lasting effects on CB morphology and function remains to be determined. Besides type I cell proliferation, other cellular components in the CB undergo proliferation and growth as well. In the nearby petrosal ganglion and superior cervical ganglion, both involved in CB chemoreception, cellular proliferation is limited to glial cells and no proliferation of neurons is observed. Also, expression of neurotrophic factors, particularly, BDNF and GDNF, is observed in type I cells of neonatal rats. Taken together, the CB undergoes significant morphological and functional changes during the postnatal period over at least 1 month. This process can be altered by oxygen concentration in ambient air.

Life-long impairment of hypoxic phrenic responses in rats following 1 month of developmental hyperoxia

Hypoxic ventilatory and phrenic responses are reduced in adult rats (3–5 months old) exposed to hyperoxia for the first month of life (hyperoxia treated). We previously reported that hypoxic phrenic responses were normal in a small sample of 14‐ to 15‐month‐old hyperoxia‐treated rats, suggesting slow, spontaneous recovery. Subsequent attempts to identify the mechanism(s) underlying this spontaneous recovery of hypoxic phrenic responses led us to re‐evaluate our earlier conclusion. Experiments were conducted in two groups of aged Sprague‐Dawley rats (14–15 months old) which were anaesthetized, vagotomized, neuromuscularly blocked and ventilated: (1) a hyperoxia‐treated group raised in 60 % O2 for the first 28 postnatal days; and (2) an age‐matched control group raised in normoxia. Increases in minute phrenic activity and integrated phrenic nerve amplitude (∫Phr) during isocapnic hypoxia (arterial partial pressures of O2, 60, 50 and 40 ± 1 mmHg) were greater in aged control (n= 15) than hyperoxia‐treated rats (n= 11; P⩽ 0.01). Phrenic burst frequency during hypoxia was not different between groups. To examine the central integration of carotid chemoafferent inputs, steady‐state relationships between carotid sinus nerve (electrical) stimulation frequency and phrenic nerve activity were compared in aged control (n= 7) and hyperoxia‐treated rats (n= 7). Minute phrenic activity, ∫Phr and burst frequency were not different between groups at any stimulation frequency between 0.5 and 20 Hz. Carotid body chemoreceptor function was examined by recording whole carotid sinus nerve responses to cessation of ventilation or injection of cyanide in aged control and hyperoxia‐treated rats. Electrical activity of the carotid sinus nerve did not change in five out of five hyperoxia‐treated rats in response to stimuli that evoked robust increases in carotid sinus nerve activity in five out of five control rats. Estimates of carotid body volume were lower in aged hyperoxia‐treated rats (4.4 (± 0.2) × 106μm3) compared to controls (17.4 (± 1.6) × 106μm3; P <0.01). We conclude that exposure to hyperoxia for the first month of life causes life‐long impairment of carotid chemoreceptor function and, consequently, blunted phrenic responses to hypoxia.

Depression of ventilation by dopamine in goats — effects of carotid body excision

Dopamine (DA) given IV by bolus injection (5, 10, 20 micrograms/kg) and by slow IV infusion (20 micrograms . kg . min) depressed VE significantly in awake normoxic goats. These responses were attenuated but not eliminated during hypoxia (FIO2 = 0.14) and hyperoxia (FIO2 = 1.0). After administering haloperidol (0.3 mg/kg) or removing the carotid bodies (CBE) there was greater attenuation of the response to DA. In normal goats haloperidol also caused a significant increase in ventilatory response to acute hypoxia and exaggerated depression of VE after 3--5 breaths O2 during steady-state hypoxia. After CBE haloperidol caused mild hypoventilation (delta PaCO2 = +2.5 Torr). CBE induced hypoventilation in goats (delta PaCO2 = +7.8 Torr) and reduced, but did not totally eliminate, peripheral chemoreceptor responses to acute stimuli (NaCN injection, transient N2 and transient O2 breathing). Attempted aortic body denervation did not eliminate these residual responses. We conclude: (1) DA may function as a modulator of carotid body (CB) function in the goat, (2) there may be central excitatory DA receptors in the goat, (3) the CB is important in regulating resting ventilation in the goat.

Carotid body hypercapnia does not elicit ventilatory acclimatization in goats

The carotid body (CB) perfusion model utilizes surgical vascular ligations to allow isolated blood supply to a single in situ CB in awake goats. The contralateral CB was excised. By use of an extracorporeal pump-oxygenator system the blood gas composition perfusing the CB can be controlled independently from that of the systemic arterial system including the brain. Using this model we compared the responses of systemically normoxic goats to CB hypercapnia and CB hypoxia. In 6 goats CB stimulation with hypercapnic-normoxic blood (mean PcbCO2 = 78 Torr, mean PcbO2 congruent to 100 Torr) produced acute hyperventilation (mean decrease in PaCO2 of 5.2 Torr, P less than 0.05) which remained constant over the 4-h perfusion period. Lack of a progressively increasing hyperventilation indicates that ventilatory acclimatization did not occur with hypercapnic CB perfusion. Hypoxic-normocapnic CB stimulation (mean PcbO2 = 40 Torr, mean PcbCO2 = 39 Torr) produced an acute mean decrease in PaCO2 of 5.5 Torr (P less than 0.05) in 6 additional goats. In contrast to CB hypercapnia, the acute hyperventilation induced by CB hypoxia was followed by a progressive time-dependent additional mean decrease in PaCO2 of 5.6 Torr (P less than 0.05) over a 4-h period (ventilatory acclimatization). These data are compatible with the concept of separate receptor mechanisms for hypercapnia and hypoxia in the CB and suggest that the early phase of ventilatory acclimatization to hypoxia in goats may result from a time-dependent increase in CB afferent output.

Lack of long-term facilitation of ventilation after exposure to hypoxia in goats.

Episodic hypoxia has been shown to induce augmented normoxic ventilatory drive or long-term facilitation (LTF, continued hyperventilation after termination of hypoxic stimulation) in awake dogs and awake goats. The main objective of these experiments was to examine whether continuous isocapnic hypoxia in awake goats elicits LTF and additionally, to determine if goats exhibit hypoxic ventilatory decline (roll-off) during the hypoxic exposure. Goats were exposed to either 4 h of isocapnic hypoxia (n = 10) or 30 min of isocapnic hypoxia (n = 7). Ventilation (VE), tidal volume and frequency were measured before, during and following the end of the isocapnic hypoxia (PaO2 40 Torr) exposure. During the 4 h period of hypoxia, VE increased in a time-dependent manner in a typical pattern of acclimatization, reaching a mean of 40.8 +/- 3.6 L/min at the end of 4 h. Five minutes after return to normoxia, VE was 13.0 +/- 0.8 L/min, not different than control VE (13.1 +/- 0.9 L/min) measured prior to the hypoxic exposure and remained unchanged from this value for another 30 min. During the 30 min hypoxic exposure, VE increased upon exposure to hypoxia, remained significantly elevated throughout the hypoxic exposure, but promptly returned to control levels upon return to normoxia. These results indicate that continuous isocapnic hypoxia elicits neither long term facilitation of ventilation nor hypoxic ventilatory decline in awake goats.

Effects of specific carotid body and brain hypoxia on respiratory muscle control in the awake goat.

1. We assessed the effects of specific brain hypoxia on the control of inspiratory and expiratory muscle electromyographic (EMG) activities in response to specific carotid body hypoxia in seven awake goats. We used an isolated carotid body perfusion technique that permitted specific, physiological, steady‐state stimulation of the carotid bodies or maintenance of normoxia and normocapnia at the carotid bodies while varying the level of systemic, and therefore, brain oxygenation. 2. Isolated brain normocapnic hypoxia of up to 1.5 h duration increased inspired minute ventilation (VI) by means of increases in both tidal volume (VT) and respiratory frequency (fR). Electromyographic activities of both inspiratory and expiratory muscles were augmented as well. These responses were similar to those produced by low levels of whole‐body normoxic hypercapnia. We conclude that moderate levels of brain hypoxia (Pa,O2 approximately 40 mmHg) in awake goats caused a net stimulation of ventilatory motor output. 3. Hypoxic stimulation of the carotid bodies alone caused comparable increases in VT and fR, and EMG augmentation of both inspiratory and expiratory muscles whether the brain was hypoxic or normoxic. These responses were quite similar to those obtained over a wide range of whole‐body normoxic hypercapnia. We conclude that the integration of carotid body afferent information is not affected by moderate brain hypoxia in awake goats. 4. We found no evidence for an asymmetrical recruitment pattern of inspiratory vs. expiratory muscles in response to carotid body hypoxia or in response to brain hypoxia alone. 5. Our data support the concept that moderate brain hypoxia results in a net stimulation of respiratory motor output. These findings question the significance of ‘central hypoxic depression’ to the regulation of breathing under physiological levels of hypoxaemia in the awake animal.

Ventilatory effects of prolonged systemic (CNS) hypoxia in awake goats.

Hypoxia isolated to the carotid body (CB) can induce time-dependent progressive hyperventilation (ventilatory acclimatization) in the absence of brain hypoxia. The studies reported in this paper were designed to determine if CNS hypoxia in the absence of CB hypoxia would affect ventilation over a 4 h period. In addition, the effect of 4 h of CNS hypoxia on the ventilatory responses to central chemoreceptor stimulation and to isolated CB stimulation were also determined. The studies were carried out in awake goats with CB blood gases controlled by an extracorporeal circuit while systemic (CNS) blood gases were determined independently by the level of inhaled gases. Systemic arterial PO2 was reduced to 40 Torr while the CB was maintained normoxic and normocapnic. Systemic arterial PCO2 was kept isocapnic. The data obtained indicate that 4 h of CNS hypoxia produced mild hyperventilation that reached a peak after 30 min of hypoxia and was sustained for the entire period of hypoxia. There was no evidence of a time-dependent progressive hyperventilation, i.e. no acclimatization. In contrast to studies in which whole body hypoxia is induced, CNS hypoxia did not result in any changes in the ventilatory responses to either central or peripheral chemoreceptor stimulation after return to normoxic conditions. These findings suggest no significant role for CNS mechanisms induced by hypoxia in ventilatory acclimatization to hypoxia in goats.

5-HT5a receptors in the carotid body chemoreception pathway of rat.

By using a specific antibody, 5-HT5a receptor-like immunoreactivity was revealed in the chemoreceptive, oxygen sensitive, carotid body (CB) type I cells, and neurons of the petrosal ganglion (PG) and the superior cervical ganglion (SCG) in rat. mRNA encoding for the 5-HTa receptor was also detected in these tissues by RT-PCR, and confirmed with DNA sequencing. The present study provides direct evidence that 5-HT5a receptors are expressed in the CB, PG and SCG, which all likely play fundamental roles in arterial chemoreception.

Domperidone-induced potentiation of ventilatory responses in awake goats

Dopamine has been implicated in maintaining tonic inhibition of carotid body activity. We tested this hypothesis by assessing the ventilatory effects of a peripheral dopamine antagonist, domperidone. The effects of this agent on the ventilatory responses to hypoxia and hypercapnia were also examined. The study was performed in awake carotid body intact and carotid body denervated goats. Resting minute ventilation increased while PaCO2 decreased (4 Torr) following domperidone administration (0.5 mg/kg, I.V.) in carotid body intact goats. This response did not occur in carotid body denervated goats supporting the hypothesis that endogenous dopamine provides tonic inhibition in the carotid body. Hypoxic and hypercapnic ventilatory responses were significantly augmented following domperidone administration in the carotid body intact goats. This supports the concept of dopaminergic modulation of the response of the carotid body to stimuli. Domperidone allows study of carotid chemoreceptor dopaminergic activity in awake animals because of its high affinity for carotid body D2 dopamine receptors and its lack of CNS effects.