Long-term effects on cardiorespiratory and behavioral responses in male and female rats prenatally exposed to cannabinoid.

Development of the respiratory system can be affected by the use of drugs during pregnancy, as the prenatal phase is highly sensitive to pharmacological interventions, resulting in long-term consequences. The deleterious effects of external cannabinoids during gestation may be related to negative interference in central nervous system formation, cardiorespiratory system function, and behavioral disorders. Nevertheless, the impact of external cannabinoids on cardiorespiratory network development, chemosensitivity, and its future consequences in adulthood is still unclear. We evaluated the effects of prenatal exposure to a synthetic cannabinoid (WIN 55,212-2, 0.5 mg.kg-1.day-1) on the cardiorespiratory control and panic-like behavior of male and female rats in adulthood. Exogenous cannabinoid exposure during pregnancy resulted in a sex-dependent difference in breathing control. Specifically, males showed increased chemosensitivity to CO2 and O2, while females exhibited decreased sensitivity. Altered cardiovascular control was evident, with prenatally treated males and females being more susceptible to hypertension and tachycardia under adverse environmental conditions. Moreover, WIN-treated males exhibited higher fragmentation of sleep episodes, while females displayed anxiolytic and panicolytic behavioral responses to CO2. However, no changes were observed in the mechanical component of the respiratory system, and there were no neuroanatomical alterations, such as changes in the expression of CB1 receptors in the brainstem or in the quantification of catecholaminergic and serotonergic neurons. These findings highlight that external interference in cannabinoid signaling during fetal development causes sex-specific long-lasting effects for the cardiorespiratory system and behavioral responses in adulthood.

Sexually dimorphic effects of prenatal diazepam exposure on respiratory control and the monoaminergic system of neonate and young rats.

Pregnancy is highly affected by anxiety disorders, which may be treated with benzodiazepines, especially diazepam (DZP), that can cross the placental barrier and interact with the fetal GABAergic system. We tested whether prenatal exposure to DZP promotes sex-specific postnatal changes in the respiratory control of rats. We evaluated ventilation ([Formula: see text]) and oxygen consumption ([Formula: see text] O2) in resting conditions and under hypercapnia (7% CO2) and hypoxia (10% O2) in newborn [postnatal day (P) 0-1 and P12-13)] and young (P21-22) rats from mothers treated with DZP during pregnancy. We also analyzed brainstem monoamines at the same ages. DZP exposure had minimal effects on room air-breathing variables in females, but caused hypoventilation (drop in [Formula: see text]/[Formula: see text] O2) in P12-13 males, lasting until P21-22. The hypercapnic ventilatory response was attenuated in P0-1 and P12-13 DZP-treated females mainly by a decrease in tidal volume (VT), whereas males had a reduction in respiratory frequency (fR) at P12-13. Minor changes were observed in hypoxia, but an attenuation in [Formula: see text] was seen in P12-13 males. In the female brainstem, DZP increased dopamine concentration and decreased 5-hydroxyindole-3-acetic acid (5-HIAA) and the 3,4-dihydroxyphenylacetic acid (DOPAC)/dopamine ratio at P0-1, and reduced DOPAC concentration at P12-13. In males, DZP decreased brainstem noradrenaline at P0-1. Our results demonstrate that prenatal DZP exposure reduces CO2 chemoreflex only in postnatal females and does not affect hypoxia-induced hyperventilation in both sexes. In addition, prenatal DZP alters brainstem monoamine concentrations throughout development differently in male and female rats.

Melanin-concentrating hormone regulates the hypercapnic chemoreflex by acting in the lateral hypothalamic area.

NEW FINDINGS: What is the central question of this study? Melanin-concentrating hormone (MCH) suppresses the hypercapnic chemoreflex: what is the mechanism by which this effect is produced? What is the main finding and its importance? MCH acting in the lateral hypothalamic area but not in the locus coeruleus in rats, in the light period, attenuates the hypercapnic chemoreflex. The data provide new insight into the role of MCH in the modulation of the hypercapnic ventilatory response. ABSTRACT: Melanin-concentrating hormone (MCH) is a hypothalamic neuropeptide involved in a broad range of homeostatic functions including regulation of the hypercapnic chemoreflex. We evaluated whether MCH modulates the hypercapnic ventilatory response by acting in the lateral hypothalamic area (LHA) and/or in the locus coeruleus (LC). Here, we measured pulmonary ventilation ( V ̇ E ${\dot V_{\rm{E}}}$ ), body temperature, electroencephalogram (EEG) and electromyogram (EMG) of unanaesthetized adult male Wistar rats before and after microinjection of MCH (0.4 mM) or MCH receptor 1 (MCH1-R) antagonist (SNAP-94847; 63 mM) into the LHA and LC, in room air and 7% CO2 conditions during wakefulness and sleep in the dark and light periods. MCH intra-LHA caused a decreased CO2 ventilatory response during wakefulness and sleep in the light period, while SNAP-94847 intra-LHA increased this response, during wakefulness in the light period. In the LC, MCH or the MCH1-R antagonist caused no change in the hypercapnic ventilatory response. Our results suggest that MCH, in the LHA, exerts an inhibitory modulation of the hypercapnic ventilatory response during the light-inactive period in rats.

Metabolic and respiratory chemosensitivity and brain monoaminergic responses to cold exposure in chicks subjected to thermal manipulation during incubation.

We evaluated ventilation (V˙E), body temperature (TB), oxygen consumption (V˙ O2), respiratory equivalent (V˙E/ V˙ O2), and monoamine concentrations of 14-day-old (14d) male and female chicks from eggs incubated at low (LT, 36 °C), control (CT, 37.5 °C) and high (HT, 39 °C) temperature during the early embryonic phase, to normoxia, hypercapnia and hypoxia under exposure to cold environment (20 °C). At normoxia, acute cold exposure did not affect the ventilatory variables, with the exception of HT males, in which cold prevented the reduced V˙E observed under thermoneutral conditions. Exposure to 20 °C caused a decrease in TB in both sexes, and LT and HT females presented a greater hypothermic response. Hypercapnia combined with cold did not alter the ventilatory variables, but LT females and CT males and females showed a blunted CO2-induced hyperventilation due to a higher V˙ O2, compared to the same groups in thermoneutral conditions. Unlike with thermoneutral conditions, the blunted hypercapnic hyperventilation observed in the HT groups was not observed during cold challenge. CO2 exposure promoted a similar decrease in TB in the thermoneutral and acutely cold exposed groups, while LT females under cold condition presented a blunted hypothermic response. During hypoxia, cold challenge attenuated the increase in V˙E in LT females and HT males, due to changes in VT. Hypoxic metabolic depression was greater in LT females and males and HT males during cold exposure, while no change in V˙E/ V˙ O2 was observed. The only alteration in monoaminergic concentration under cold challenge was an increase in brainstem 5-HIAA and 5-HIAA/5-HT ratio in HT females, and an enhanced 5-HT concentration in HT males. In summary, thermal manipulation during embryogenesis induces 14d old chicks to respond differently to cold stress with LT females and HT males being more sensitive.

Sex differences in breathing.

Breathing is a vital behavior that ensures both the adequate supply of oxygen and the elimination of CO2, and it is influenced by many factors. Despite that most of the studies in respiratory physiology rely heavily on male subjects, there is much evidence to suggest that sex is an important factor in the respiratory control system, including the susceptibility for some diseases. These different respiratory responses in males and females may be related to the actions of sex hormones, especially in adulthood. These hormones affect neuromodulatory systems that influence the central medullary rhythm/pontine pattern generator and integrator, sensory inputs to the integrator and motor output to the respiratory muscles. In this article, we will first review the sex dependence on the prevalence of some respiratory-related diseases. Then, we will discuss the role of sex and gonadal hormones in respiratory control under resting conditions and during respiratory challenges, such as hypoxia and hypercapnia, and whether hormonal fluctuations during the estrous/menstrual cycle affect breathing control. We will then discuss the role of the locus coeruleus, a sexually dimorphic CO2/pH-chemosensitive nucleus, on breathing regulation in males and females. Next, we will highlight the studies that exist regarding sex differences in respiratory control during development. Finally, the few existing studies regarding the influence of sex on breathing control in non-mammalian vertebrates will be discussed.

Brainstem catecholaminergic neurones and breathing control during postnatal development in male and female rats.

KEY POINTS: The brainstem catecholaminergic (CA) modulation on ventilation changes with development. We determined the role of the brainstem CA system in ventilatory control under normocapnic and hypercapnic conditions during different phases of development [postnatal day (P)7-8, P14-15 and P20-21] in male and female Wistar rats. Brainstem CA neurones produce a tonic inhibitory drive that affects breathing frequency in P7-8 rats and provide an inhibitory drive during hypercapnic conditions in both males and females at P7-8 and P14-15. In pre-pubertal rats, brainstem CA neurones become excitatory for the CO2 ventilatory response in males but remain inhibitory in females. Diseases such as sudden infant death syndrome, congenital central hypoventilation syndrome and Rett syndrome have been associated with abnormalities in the functioning of CA neurones; therefore, the results of the present study contribute to a better understanding of this system. ABSTRACT: The respiratory network undergoes significant development during the postnatal phase, including the maturation of the catecholaminergic (CA) system. However, postnatal development of this network and its effect on the control of pulmonary ventilation ( V̇E ) is not fully understood. We investigated the involvement of brainstem CA neurones in respiratory control during postnatal development [postnatal day (P)7-8, P14-15 and P20-21], in male and female rats, through chemical injury with conjugated saporin anti-dopamine ß-hydroxylase (DßH-SAP). Thus, DßH-SAP (420 ng µL-1 ), saporin (SAP) or phosphate buffered solution (PBS) was injected into the fourth ventricle of neonatal Wistar rats of both sexes. V̇E and oxygen consumption were recorded 1 week after the injections in unanaesthetized neonatal and juvenile rats during room air and hypercapnia. The resting ventilation was higher in both male and female P7-8 lesioned rats by 33%, with a decrease in respiratory variability being observed in males. The hypercapnic ventilatory response (HCVR) was altered in male and female lesioned rats at all postnatal ages. At P7-8, the HCVR for males and females was increased by 37% and 30%, respectively. For both sexes at P14-15 rats, the increase in V̇E during hypercapnia was 37% higher for lesioned rats. A sex-specific difference in HCRV was observed at P20-21, with lesioned males showing a 33% decrease, and lesioned females showing an increase of 33%. We conclude that brainstem CA neurones exert a tonic inhibitory effect on V̇E in the early postnatal days of the life of a rat, increase variability in P7-8 males and modulate HCRV during the postnatal phase.

Exercise derived myokine irisin as mediator of cardiorespiratory, metabolic and thermal adjustments during central and peripheral chemoreflex activation.

Exercise elicits physiological adaptations, including hyperpnea. However, the mechanisms underlying exercise-induced hyperpnea remain unresolved. Skeletal muscle acts as a secretory organ, releasing irisin (IR) during exercise. Irisin can cross the blood-brain barrier, influencing muscle and tissue metabolism, as well as signaling in the central nervous system (CNS). We evaluated the effect of intracerebroventricular or intraperitoneal injection of IR in adult male rats on the cardiorespiratory and metabolic function during sleep-wake cycle under room air, hypercapnia and hypoxia. Central IR injection caused an inhibition on ventilation (VE) during wakefulness under normoxia, while peripheral IR reduced VE during sleep. Additionally, central IR exacerbates hypercapnic hyperventilation by increasing VE and reducing oxygen consumption. As to cardiovascular regulation, central IR caused an increase in heart rate (HR) across all conditions, while no change was observed following peripheral administration. Finally, central IR attenuated the hypoxia-induced regulated hypothermia and increase sleep episodes, while peripheral IR augmented CO2-induced hypothermia, during wakefulness. Overall, our results suggest that IR act mostly on CNS exerting an inhibitory effect on breathing under resting conditions, while stimulating the hypercapnic ventilatory response and increasing HR. Therefore, IR seems not to be responsible for the exercise-induced hyperpnea, but contributes to the increase in HR.

Locus coeruleus noradrenaline depletion and its differential impact on CO2-induced panic and hyperventilation in male and female mice.

CO2 exposure has been used to investigate the panicogenic response in patients with panic disorder. These patients are more sensitive to CO2, and more likely to experience the "false suffocation alarm" which triggers panic attacks. Imbalances in locus coeruleus noradrenergic (LC-NA) neurotransmission are responsible for psychiatric disorders, including panic disorder. These neurons are sensitive to changes in CO2/pH. Therefore, we investigated if LC-NA neurons are differentially activated after severe hypercapnia in mice. Further, we evaluated the participation of LC-NA neurons in ventilatory and panic-like escape responses induced by 20% CO2 in male and female wild type mice and two mouse models of altered LC-NA synthesis. Hypercapnia activates the LC-NA neurons, with males presenting a heightened level of activation. Mutant males lacking or with reduced LC-NA synthesis showed hypoventilation, while animals lacking LC noradrenaline present an increased metabolic rate compared to wild type in normocapnia. When exposed to CO2, males lacking LC noradrenaline showed a lower respiratory frequency compared to control animals. On the other hand, females lacking LC noradrenaline presented a higher tidal volume. Nevertheless, no change in ventilation was observed in either sex. CO2 evoked an active escape response. Mice lacking LC noradrenaline had a blunted jumping response and an increased freezing duration compared to the other groups. They also presented fewer racing episodes compared to wild type animals, but not different from mice with reduced LC noradrenaline. These findings suggest that LC-NA has an important role in ventilatory and panic-like escape responses elicited by CO2 exposure in mice.

Intra-uterine diazepam exposure decreases the number of catecholaminergic and serotoninergic neurons of neonate rats.

Benzodiazepines, such as diazepam (DZP), are used to treat anxiety disorders, and are prescribed to pregnant woman for therapeutic purposes. Concerns regarding their consequences on postnatal development rise as they cross the placenta and interact with the embryo. Occurrence of malformation and behavioral syndromes have been reported for different ages, but little is known about their effects on the brain after exposure during intrauterine life. Thus, we sought to evaluate the effects of intrauterine exposure to DZP on the number of brainstem's catecholaminergic and serotonergic neurons, implicated in respiratory control, in male and female rats on postnatal (P) day 12-13, using immunofluorescence labeling for tyrosine-hydroxylase (TH) and serotonin (5-HT). We observed a reduction in the number of catecholaminergic neurons for males and females. Special attention is given to the reduction in the density of neurons in the A6 region, involved in ventilatory responses to CO2. Interestingly, only males showed a reduction in the number of serotonergic neurons, while females were not affected. These findings suggest that in utero exposure to DZP results in deleterious neuroanatomical effects on P12-13 rats and raises a note of concern for women clinicians to make more informed choices about the use of anxiolytic treatments during gestation.

Heat stress affects breathing and metabolism of chicks incubated at high temperature.

Global warming poses serious implications to animal physiology and a gradual increase in ambient temperature affects all living organisms, particularly fast-growing selected species. We recorded ventilation (V̇E), body temperature (TB), oxygen consumption (V̇O2) and respiratory equivalent (V̇E/V̇O2) of 14-day-old (14d) male and female chicks at room air conditions, hypercapnia and hypoxia at heat stress (HS, 32 °C). These chicks had previously been exposed to control (CI, 37.5 °C) and high (HI, 39 °C) temperatures during the first 5 days of incubation. Under resting conditions, acute HS increased V̇E in HI females but not in HI males. Hypercapnia combined with heat promoted a potentiation of CO2-hyperventilatory response in HI females when compared with thermoneutral condition, whereas in HI incubated males a hypoventilation under hypercapnia and heat stress was observed compared to the CI group. Hypoxia associated with heat stress increased V̇E only in HI females. Our data indicates that females are more sensitive to thermal manipulation during incubation and it seems that the thermal embryonic manipulation, at least during the first days of development, does not improve the adaptive response of chicks to heat stress.

Sex- and age-specific respiratory alterations induced by prenatal exposure to the cannabinoid receptor agonist WIN 55,212-2 in rats.

BACKGROUND AND PURPOSE: Cannabis legalization has risen in many countries, and its use during pregnancy has increased. The endocannabinoid system is present in the CNS at early stages of embryonic development, and regulates functional brain maturation including areas responsible for respiratory control, data on the influence of external cannabinoids on the development of the respiratory system and possible consequences during postnatal life are limited. EXPERIMENTAL APPROACH: We evaluated the effects of prenatal exposure to synthetic cannabinoid (WIN 55,212-2 [WIN], 0.5 mg·kg-1 ·day-1 ) on the respiratory control system in neonatal (P0, P6-7 and P12-13) and juvenile (P27-28) male and female rats. KEY RESULTS: WIN administration to pregnant rats interfered sex-specifically with breathing regulation of offspring, promoting a greater sensitivity to CO2 at all ages in males (except P6-7) and in juvenile females. An altered hypoxic chemoreflex was observed in P0 (hyperventilation) and P6-7 (hypoventilation) males, which was absent in females. Along with breathing alterations, brainstem analysis showed an increase in the number of catecholaminergic neurons and cannabinoid receptor type 1 (CB1 ) and changes in tissue respiration in the early males. A reduction in pulmonary compliance was observed in juvenile male rats. Preexposure to WIN enhanced spontaneous apnoea and reduced the number of serotoninergic (5-HT) neurons in the raphe magnus nucleus of P0 females. CONCLUSIONS AND IMPLICATIONS: These data demonstrate that excess stimulation of the endocannabinoid system during gestation has prolonged and sex-specific consequences for the respiratory control system.

Inhibition of Pro-Inflammatory Microglia with Minocycline Improves Cognitive and Sleep-Wake Dysfunction Under Respiratory Stress in a Sporadic Model for Alzheimer's Disease.

BACKGROUND: Neuroinflammation in Alzheimer's disease (AD) can occur due to excessive activation of microglia in response to the accumulation of amyloid-ß peptide (Aß). Previously, we demonstrated an increased expression of this peptide in the locus coeruleus (LC) in a sporadic model for AD (streptozotocin, STZ; 2 mg/kg, ICV). We hypothesized that the STZ-AD model exhibits neuroinflammation, and treatment with an inhibitor of microglia (minocycline) can reverse the cognitive, respiratory, sleep, and molecular disorders of this model. OBJECTIVE: To evaluate the effect of minocycline treatment in STZ model disorders. METHODS: We treated control and STZ-treated rats for five days with minocycline (30 mg/kg, IP) and evaluated cognitive performance, chemoreflex response to hypercapnia and hypoxia, and total sleep time. Additionally, quantification of Aß, microglia analyses, and relative expression of cytokines in the LC were performed. RESULTS: Minocycline treatment improved learning and memory, which was concomitant with a decrease in microglial cell density and re-establishment of morphological changes induced by STZ in the LC region. Minocycline did not reverse the STZ-induced increase in CO2 sensitivity during wakefulness. However, it restored the daytime sleep-wake cycle in STZ-treated animals to the same levels as those observed in control animals. In the LC, levels of A and expression of Il10, Il1b, and Mcp1 mRNA remained unaffected by minocycline, but we found a strong trend of minocycline effect on Tnf- α. CONCLUSION: Our findings suggest that minocycline effectively reduces microglial recruitment and the inflammatory morphological profile in the LC, while it recovers cognitive performance and restores the sleep-wake pattern impaired by STZ.

Prenatal fluoxetine has long-lasting, differential effects on respiratory control in male and female rats.

Serotonin (5-HT) is an important modulator of brain networks that control breathing. The selective serotonin reuptake inhibitor fluoxetine (FLX) is the first-line antidepressant drug prescribed during pregnancy. We investigated the effects of prenatal FLX exposure on baseline breathing, ventilatory and metabolic responses to hypercapnia and hypoxia as well as number of brainstem 5-HT and tyrosine hydroxylase (TH) neurons of rats during postnatal development (P0-82). Prenatal FLX exposure of males showed a lower baseline V̇e that appeared in juveniles and remained in adulthood, with no sleep-wake state dependency. Prenatal FLX exposure of females did not affect baseline breathing. Juvenile male FLX showed increased CO2 and hypoxic ventilatory responses, normalizing by adulthood. Alterations in juvenile FLX-treated males were associated with a greater number of 5-HT neurons in the raphe obscurus (ROB) and raphe magnus (RMAG). Adult FLX-exposed males showed greater number of 5-HT neurons in the raphe pallidus (RPA) and TH neurons in the A5, whereas reduced number of TH neurons in A7. Prenatal FLX exposure of female rats was associated with greater hyperventilation induced by hypercapnia at P0-2 and juveniles, whereas P12-14 and adult FLX (non-rapid eye movement, NREM sleep) rats showed an attenuation of the hyperventilation induced by CO2. FLX-exposed females had fewer 5-HT neurons in the RPA and reduced TH A6 density at P0-2; and greater number of TH neurons in the A7 at P12-14. These data indicate that prenatal FLX exposure affects the number of some monoaminergic regions in the brain and results in long-lasting, sex-specific changes in baseline breathing pattern and ventilatory responses to respiratory challenges.NEW & NOTEWORTHY Selective serotonin reuptake inhibitors (SSRIs) readily cross the placental and the fetal blood-brain barrier where it will affect 5-HT levels in the developing brain. Although SSRI is used during pregnancy, there are no studies showing SSRI exposure during late pregnancy and postnatal effects on breathing control in males and females. We demonstrated that fluoxetine exposure during late pregnancy in rats was associated with long-lasting, sex-specific effects on breathing and brainstem monoaminergic groups.

An age- and sex-dependent role of catecholaminergic neurons in the control of breathing and hypoxic chemoreflex during postnatal development.

The respiratory system undergoes significant development during the postnatal phase. Maturation of brainstem catecholaminergic (CA) neurons is important for the control and modulation of respiratory rhythmogenesis, as well as for chemoreception in early life. We demonstrated an inhibitory role for CA neurons in CO2 chemosensitivity in neonatal and juvenile male and female rats, but information regarding their role in the hypoxic ventilatory response (HVR) is lacking. We evaluated the contribution of brainstem CA neurons in the HVR during postnatal (P) development (P7-8, P14-15 and P20-21) in male and female rats through chemical injury with conjugated saporin anti-dopamine beta-hydroxylase (DßH-SAP, 420 ng·µL-1) injected in the fourth ventricle. Ventilation (V̇E) and oxygen consumption were recorded one week after the lesion in unanesthetized rats during exposure to normoxia and hypoxia. Hypoxia reduced breathing variability in P7-8 control rats of both sexes. At P7-8, the HVR for lesioned males and females increased 27% and 24%, respectively. Additionally, the lesion reduced the normoxic breathing variability in both sexes at P7-8, but hypoxia partially reverted this effect. For P14-15, the increase in V̇E during hypoxia was 30% higher for male and 24% higher for female lesioned animals. A sex-specific difference was detected at P20-21, as lesioned males exhibited a 24% decrease in the HVR, while lesioned females experienced a 22% increase. Furthermore, the hypoxia-induced body temperature reduction was attenuated in P20-21 lesioned females. We conclude that brainstem CA neurons modulate the HRV during the postnatal phase, and possibly thermoregulation during hypoxia.

Functional role for preoptic CB1 receptors in breathing and thermal control.

The anteroventral preoptic region (AVPO) of the hypothalamus is involved in both temperature and breathing regulation. This area densely express cannabinoid receptors type 1 (CB1) that modulate both excitatory and inhibitory synaptic transmission. However, it is still unknown if the endocannabinoid system located in the AVPO participates in breathing control and thermoregulation. Therefore, we tested the participation of CB1 in the AVPO in the modulation of ventilation and thermal control during normoxia and hypoxia. To this end, body temperature (Tb) of Wistar rats was monitored by datallogers and ventilation (VE) by whole body plethysmography before and after intra-AVPO microinjection of AM-251 (CB1 antagonist, 50 and 100 pmol) followed by 60 min of hypoxia exposure (7% O2). Intra-AVPO microinjection of the higher dose of AM-251 increased VE but did not change Tb under resting conditions. Exposure of rats to 7% of inspired oxygen evoked typical hypoxia-induced anapyrexia and hyperventilation after vehicle microinjection. The higher dose of the cannabinoid antagonist increased the hypoxia-induced hyperventilation, in the same magnitude as observed under normoxic condition, whereas the drop in Tb elicited by hypoxia was attenuated. Therefore, the present results demonstrate that the endocannabinoid system acting on CB1 receptors in the AVPO exerts a tonic inhibitory modulation on breathing but seem not be involved in thermoregulation during resting conditions. In addition, activation of CB1 receptors in the AVPO stimulate thermal response during hypoxia, reducing energetically expensive responses, such as the hypoxic hyperventilation.

The role of testosterone in the respiratory and thermal responses to hypoxia and hypercapnia in rats.

Many diseases of the respiratory system occur differently in males and females, indicating a possible role of gonadal hormones in respiratory control. We hypothesized that testosterone (T) is important for the ventilatory chemosensitivity responses in males. To test this hypothesis, we evaluated ventilation (V̇ E), metabolic rate and body temperature (Tb) under normoxia/normocapnia, hypercapnia and hypoxia in orchiectomized (ORX), ORX with testosterone replacement (ORX + T) or flutamide (FL, androgen receptor blocker)-treated rats. We also performed immunohistochemistry to evaluate the presence of androgen receptor (AR) in the carotid body (CB) of intact males. Orchiectomy promoted a reduction V̇ E and ventilatory equivalent (V̇ E /V̇ O2) under room-air conditions, which was restored with testosterone treatment. Moreover, during hypoxia or hypercapnia, animals that received testosterone replacement had a higher V̇ E and V̇ E /V̇ O2 than control and ORX, without changes in metabolic and thermal variables. Flutamide decreased the hypoxic ventilatory response without changing the CO2-drive to breathe, suggesting that the testosterone effect on hypercapnic hyperventilation does not appear to involve the AR. We also determined the presence of AR in the CB of intact animals. Our findings demonstrate that testosterone seems to be important for maintaining resting V̇ E in males. In addition, the influence of testosterone on V̇ E, either during resting conditions or under hypoxia and hypercapnia, seems to be a direct and specific effect, as no changes in metabolic rate or Tb were observed during any treatment. Finally, a putative site of testosterone action during hypoxia is the CB, since we detected the presence of AR in this structure.

TRPV1 Inhibits the Ventilatory Response to Hypoxia in Adult Rats, but Not the CO2-Drive to Breathe.

Receptors of the transient receptor potential (TRP) channels superfamily are expressed in many tissues and have different physiological functions. However, there are few studies investigating the role of these channels in cardiorespiratory control in mammals. We assessed the role of central and peripheral TRPV1 receptors in the cardiorespiratory responses to hypoxia (10% O2) and hypercapnia (7% CO2) by measuring pulmonary ventilation ( V ˙ E ), heart rate (HR), mean arterial pressure (MAP) and body temperature (Tb) of male Wistar rats before and after intraperitoneal (AMG9810 [2.85 µg/kg, 1 mL/kg]) or intracebroventricular (AMG9810 [2.85 µg/kg, 1 µL] or AMG7905 [28.5 µg/kg, 1 µL]) injections of TRPV1 antagonists. Central or peripheral injection of TRPV1 antagonists did not change cardiorespiratory parameters or Tb during room air and hypercapnic conditions. However, the hypoxic ventilatory response was exaggerated by both central and peripheral injection of AMG9810. In addition, the peripheral antagonist blunted the drop in Tb induced by hypoxia. Therefore, the current data provide evidence that TRPV1 channels exert an inhibitory modulation on the hypoxic drive to breathe and stimulate the Tb reduction during hypoxia.

Participation of locus coeruleus in breathing control in female rats.

Several evidences indicate that the locus coeruleus (LC) is involved in central chemoreception responding to CO2/pH and displaying a high percentage of chemosensitive neurons (>80%). However, there are no studies about the LC-mediated hypercapnic ventilation performed in females. Therefore, we assessed the role of noradrenergic LC neurons in non-ovariectomized (NOVX), ovariectomized (OVX) and estradiol (E2)-treated ovariectomized (OVX+E2) rats in respiratory response to hypercapnia, using a 6-hydroxydopamine (6-OHDA) - lesion model. A reduction in the number of tyrosine hydroxylase (TH) immunoreactive neurons (51-90% in 3 animals of NOVX group, 20-42% of lesion in 5 animals of NOVX females, 61.3% for OVX and 62.6% for OVX+E2 group) was observed seven days after microinjection of 6-OHDA in the LC. The chemical lesion of the LC resulted in decreased respiratory frequency under normocapnic conditions in OVX and OVX+E2 group. Hypercapnia increased ventilation in all groups as consequence of increases in respiratory frequency (fR) and tidal volume (VT). Nevertheless, the hypercapnic ventilatory response was significantly decreased in 6-OHDA-NOVX>50% rats compared with SHAM-NOVX group and with females that had 20-42% of LC lesion. In OVX and OVX+E2 lesioned groups, no difference in CO2 ventilatory response was observed when compared to SHAM-OVX and SHAM-OVX+E2 groups, respectively. Neither basal body temperature (Tb) nor Tb reduction in response to hypercapnia were affected by E2 treatment, ovariectomy or LC lesion. Thus, our data show that LC noradrenergic neurons seem to exert an excitatory role on the hypercapnic ventilatory response in female rats, as evidenced by the results in NOVX animals with LC lesioned more than 50%; however, this modulation is not observed in OVX and OVX+E2 rats. In addition, LC noradrenergic neurons of OVX females seem to provide a tonic excitatory drive to maintain breathing frequency in normocapnia, and this response may not to be functionally influenced by E2.