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.

Starch and fiber intake effects on energy metabolism, growth, and carapacial scute pyramiding of red-footed tortoise hatchlings (Chelonoidis carbonaria).

Tortoise husbandry includes reports of excessive growth and carapace pyramiding, although triggers still remain to be fully elucidated. Juvenile red-footed tortoises (Chelonoidis carbonaria) were fed with two different diets, one high in fiber (HF; 14.2% crude fiber; 39.2% neutral detergent fiber, NDF; dry matter basis, DMB) and one high in starch (HS; 27.7% DMB), to assess effects on energy metabolism, nutrient digestibility, and growth. A total of 20 hatchlings (10 per diet) were used to evaluate: apparent digestibility coefficients (Da) of nutrients and gross energy (GE), passage times at 5 and 11 months of age; resting and post-prandial metabolic rates at 6 and 12 months of age; growth rates; pyramiding; and estimated body composition. Animals fed HS showed higher mass-specific intake of digestible energy (113.9 ± 32.1 kJ kg-1 day-1 vs. 99.6 ± 35.3 kJ kg-1 day-1; P < 0.05), digestible DM (6.1 ± 1.8 g kg-1 day-1 vs. 5.0 ± 1.8 g kg-1 day-1; P < 0.01), shorter transit (3 ± 1 days vs. 4 ± 1 days; P < 0.01) and retention times (8 ± 2 days vs. 10 ± 2 days; P < 0.01), and higher Da of DM, starch, NDF, and GE. Crude protein Da was higher for HF. Rest and post-prandial metabolic rates, and pyramiding degree were not affected by diets. At 13 months, the animals from HS presented wider plastrons and carapaces, and higher carapace width growth rates. In addition, these animals had lower body mineral content (1.88 ± 0.15% vs. 2.15 ± 0.19%; P < 0.01) and bone density (0.13 ± 0.01 g mm-2 vs. 0.15 ± 0.02 g mm-2; P < 0.02). Results provide evidence that highly digestible foods can accelerate shell growth and lower mineralization in this species.

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.

A5 noradrenergic neurons and breathing control in neonate rats.

The pontine A5 noradrenergic group contributes to the maturation of the respiratory system before birth in rats. These neurons are connected to the neural network responsible for respiratory rhythmogenesis. In the present study, we investigated the participation of A5 noradrenergic neurons in neonates (P7-8 and P14-15) in the control of ventilation during hypoxia and hypercapnia in in vivo experiments using conjugated saporin anti-dopamine beta-hydroxylase (DßH-SAP) to specifically ablate noradrenergic neurons. Thus, DßH-SAP (420 ng/µL) or saporin (SAP, control) was injected into the A5 region of neonatal male Wistar rats. Hypoxia reduced respiratory variability in control animals; however, A5 lesion prevented this effect in P7-8 rats. Our data suggest that noradrenergic neurons of the A5 region in neonate rats do not participate in the control of ventilation under baseline and hypercapnic conditions, but exert an inhibitory modulation on breathing variability under hypoxic challenge in early life (P7-8).

Influence of incubation temperature on embryo development, hatchling morphology and early growth rate in red-footed tortoise (Chelonoidis carbonaria).

Reptile embryos respond to temperature changes with metabolic and physiological adjustments that influence hatchling success, phenotype, behaviour, and growth rate. Climate change and global warming can affect the reptile population by altering the frequencies of hatchling survival and phenotypes. Therefore, previous studies proposed artificial incubation as a potential strategy for mitigating these effects. Red-footed tortoise (Chelonoidis carbonaria) eggs were collected and incubated at constant temperatures of 27.5 °C and 29.5 °C to investigate the physiological effects of temperature on embryo development, hatchling morphology, and early post-hatch growth rate. The direct effect of temperature on the incubation period, egg mass loss, hatching success, hatchling size, and mass was evaluated at hatching and three months of age. Hatchlings from 29.5 °C presented a shorter incubation period (141 days) than those from 27.5 °C (201 days; p < 0.05). Egg mass loss, hatchling mass, and size at hatching were not different between the incubation temperatures (p > 0.05). However, the hatching success (survival rate) was lower (64.5% versus 100%) in eggs incubated at 29.5 °C, but the hatchling mass and straight plastron width were higher at three months of age than those from eggs incubated at 27.5 °C (p < 0.05). These results indicate that incubation temperature influences hatching success and hatchling size and mass in the first months by influencing the early growth rate.

Hormonal correlates of the annual cycle of activity and body temperature in the South-American tegu lizard (Salvator merianae).

Life history transitions and hormones are known to interact and influence many aspects of animal physiology and behavior. The South-American tegu lizard (Salvator merianae) exhibits a profound seasonal shift in metabolism and body temperature, characterized by high daily activity during warmer months, including reproductive endothermy in spring, and metabolic suppression during hibernation in winter. This makes S. merianae an interesting subject for studies of interrelationships between endocrinology and seasonal changes in physiology/behavior. We investigated how plasma concentrations of hormones involved in regulation of energy metabolism (thyroid hormones T4 and T3; corticosterone) and reproduction (testosterone in males and estrogen/progesterone in females) correlate with activity and body temperature (Tb) across the annual cycle of captive held S. merianae in semi-natural conditions. In our initial model, thyroid hormones and corticosterone showed a positive relationship with activity and Tb with independent of sex: T3 positively correlated with activity and Tb, while T4 and corticosterone correlated positively with changes in Tb only. This suggests that thyroid hormones and glucocorticoids may be involved in metabolic transitions of annual cycle events. When accounting for sex-steroid hormones, our sex separated models showed a positive relationship between testosterone and Tb in males and progesterone and activity in females. Coupling seasonal endocrine measures with activity and Tb may expand our understanding of the relationship between animal's physiology and its environment. Manipulative experiments are required in order to unveil the directionality of influences existing among abiotic factors and the hormonal signaling of annual cyclicity in physiology/behavior.

Seasonal changes in plasma concentrations of the thyroid, glucocorticoid and reproductive hormones in the tegu lizard Salvator merianae.

The tegu lizard Salvator merianae is a large, widely distributed teiid lizard endemic to South America that exhibits annual cycles of high activity during the spring and summer, and hibernation during winter. This pattern of activity and hibernation is accompanied by profound seasonal changes in physiology and behavior, including endothermy during the austral spring. The unusual combination of seasonal endothermy, hibernation and oviparity, in a non-avian, non-mammalian species, makes S. merianae an interesting subject for study of comparative aspects of endocrine regulation of seasonal changes in physiology. In the present study, we first validated commercially available immunoassay kits for quantification of hormone concentrations of the reproductive (testosterone, estradiol and progesterone), adrenal (corticosterone), and thyroid [thyroxine (T4) and triiodothyronine (T3)] axes in plasma of an outdoor, captive adult male and female S. merianae in southeastern Brazil. All assays exhibited parallelism and accuracy with S. merianae plasma. We next assessed patterns of concentration of these hormones across the annual cycle of S. merianae. Testosterone in males and estradiol in females peaked in spring coincident with the peak in reproductive behavior. Progesterone in females was significantly elevated in October coincident with putative ovulation when gravid females build nests. Thyroid hormones, known for regulating energy metabolism, varied seasonally with some sex-dependent differences. T4 gradually increased from an annual nadir during pre-hibernation and hibernation to high concentrations during spring in both sexes. In contrast, T3 did not vary seasonally in males, but females showed a two-fold increase in T3 during the spring reproductive season. T3 may be involved in energy investment during the seasonal production of large clutches of eggs. Corticosterone was significantly elevated during the active season in both sexes, suggesting its involvement in mobilization of energy stores and modulation of behavior (territoriality) and physiology. Ours is the first investigation of concurrent changes in reproductive, thyroid and adrenal hormone concentrations in this endemic and physiologically unique South American lizard. Our findings set the stage for future investigations to determine the extent to which these hormones influence activity and thermoregulation in S. merianae.

Seasonal variation of hypoxic and hypercarbic ventilatory responses in the lizard Tropidurus torquatus.

Carbon dioxide (CO2) and oxygen (O2) influence the breathing pattern of reptiles, especially when CO2 is in excess or O2 at low concentrations and the effects of these gases on the respiratory response varies according to the species. In addition to respiratory gases, seasonal changes can also modulate breathing pattern and ventilatory responses to hypoxia and hypercarbia. Therefore, the present study investigated the breathing pattern and ventilatory responses to hypercarbia (5% CO2) and hypoxia (5% O2) of the Neotropical lizard Tropidurus torquatus over a period of one year, covering all seasons (summer, autumn, winter and spring). Our data suggest that like other ectothermic sauropsids, Tropidurus torquatus possesses distinct ventilatory responses to hypoxia and hypercarbia, being more sensitive to changes in CO2 than in O2. Additionally, the ventilatory responses to hypoxia were more pronounced during summer and hypercanic and pos-hypercapnic ventilatory response was reduced during spring, suggesting that seasonality modulates the control of ventilation in this species.

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.

Mu and kappa opioid receptors of the periaqueductal gray stimulate and inhibit thermogenesis, respectively, during psychological stress in rats.

The periaqueductal gray matter (PAG) is rich in mu and kappa opioid receptors, and this system is involved in thermoregulation, analgesia, and defensive behaviors. No study approached the involvement of the PAG opioids in body temperature (Tb) regulation during psychological stress such as restraint. Because activation of mu and kappa receptors increases and reduces Tb, respectively, we tested the hypothesis that they exert excitatory and inhibitory modulation, respectively, of the restraint-induced fever in rats. To this end, Tb, heat loss index (HLI, inference for peripheral vasoconstriction/vasodilation), and oxygen consumption (inference for thermogenesis) were monitored in unanesthetized rats, restrained or unrestrained, before and after intra-PAG microinjection of the selective mu opioid receptor antagonist (D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 cyclic, CTAP; 1 and 10 µg/100 nL) or the selective kappa opioid receptor antagonist (nor-binaltorphimine dihydrochloride, nor-BNI; 1 and 4 µg/100 nL) or saline (100 nL). CTAP and nor-BNI did not change the Tb or HLI of euthermic animals. During restraint, Tb increased (1.0 ± 0.1 °C) in all groups; however, this effect was lower in those animals treated with CTAP and higher in animals treated with nor-BNI. The HLI decreased during restraint and increased after animals were released, but this response was not affected by any treatment. Restraint stress increased oxygen consumption (35.9 ± 3.9% elevation), but this response was diminished by CTAP and overstimulated by nor-BNI. Confirming our hypothesis, the results indicate that the mu and kappa opioid receptors in the PAG of rats play a pyrogenic and antipyretic role, respectively, during fever induced by restraint by affecting the thermogenic but not the heat conservation effector.

Influence of estrous cycle hormonal fluctuations and gonadal hormones on the ventilatory response to hypoxia in female rats.

Sex hormones may influence many physiological processes. Recently, we demonstrated that hormonal fluctuations of cycling female rats do not affect respiratory parameters during hypercapnia. However, it is still unclear whether sex hormones and hormonal fluctuations that occur during the estrous cycle can affect breathing during a hypoxic challenge. Our study aimed to evaluate respiratory, metabolic, and thermal responses to hypoxia in female rats on different days of the estrous cycle (proestrus, estrus, metestrus, and diestrus) and in ovariectomized rats that received replacement with oil (OVX), estradiol (OVX + E2), or a combination of estradiol and progesterone (OVX + E2P). Ventilation (V E), tidal volume (V T), respiratory frequency (fR), oxygen consumption (VO2), and V E/VO2 were not different during the estrous cycle in normoxia or hypoxia. Body temperature (Tb) was higher during estrus, but decreased similarly in all groups during hypoxia. Compared with intact females in estrus, gonadectomized rats also had lower Tb in normoxia, but not in hypoxia. OVX rats experienced a significant drop in the ventilatory response to hypoxia, but hormonal replacement did not restore values to the levels of an intact animal. Our data demonstrate that the different phases of the estrous cycle do not alter ventilation during normoxia and hypoxia, but OVX animals display lower ventilatory responses to hypoxia compared with ovary-intact rats. Because estradiol and progesterone replacement did not cause significant differences in ventilation, our findings suggest that a yet-to-be-defined non-steroidal ovarian hormone is likely to stimulate the ventilatory responses to hypoxia in females.

Thermal tachypnea in avian embryos.

Many adult mammals and birds respond to high surrounding temperatures with thermal tachypnea - an increase in breathing frequency accompanied by shallow tidal volume, with minimal increase in oxygen consumption (V̇O2 ). This pattern favors heat dissipation by evaporative water loss (EWL) through the respiratory tract. We asked to what extent this response was apparent at the earliest stages of development, when pulmonary ventilation initiates. Measurements of pulmonary ventilation (V̇E; barometric technique), V̇O2  (open-flow methodology) and EWL (water scrubbers) were performed on chicken embryos at the earliest appearance of pulmonary ventilation, during the internal pipping stage. Data were collected, first, at the normal incubation temperature (37.5°C); then, ambient and egg temperatures were increased to approximately 44°C over a 2 h period. Other embryos of the same developmental stage (controls) were maintained in normothermia for the whole duration of the experiment. During heat exposure, the embryo's V̇O2  and carbon dioxide production increased little. In contrast, V̇E more than doubled (∼128% increase), entirely because of the large rise in breathing frequency (∼132% increase), with no change in tidal volume. EWL did not change significantly, probably because, within the egg, the thermal and water vapor gradients are almost nonexistent. We conclude that chicken embryos respond to a major heat load with tachypnea, like many adult mammals and birds do. Its appearance so early in development, although ineffective for heat loss, signifies that thermal tachypnea represents an important breathing response necessary to be functional from hatching.

Temperature effects on the cardiorespiratory control of American bullfrog tadpoles based on a non-invasive methodology.

Temperature effects on cardiac autonomic tonus in amphibian larval stages have never been investigated. Therefore, we evaluated the effect of different temperatures (15, 25 and 30°C) on the cardiorespiratory rates and cardiac autonomic tonus of premetamorphic tadpoles of the bullfrog, Lithobates catesbeianus To this end, a non-invasive method was developed to permit measurements of electrocardiogram (ECG) and buccal movements (fB; surface electromyography of the buccal floor). For evaluation of autonomic regulation, intraperitoneal injections of Ringer solution (control), atropine (cholinergic muscarinic antagonist) and sotalol (ß-adrenergic antagonist) were performed. Ringer solution injections did not affect heart rate (fH) or fB across temperatures. Cardiorespiratory parameters were significantly augmented by temperature (fH: 24.5±1.0, 54.5±2.0 and 75.8±2.8 beats min-1 at 15, 25 and 30°C, respectively; fB: 30.3±1.1, 73.1±4.0 and 100.6±3.7 movements min-1 at 15, 25 and 30°C, respectively). A predominant vagal tone was observed at 15°C (32.0±3.2%) and 25°C (27.2±6.7%) relative to the adrenergic tone. At 30°C, the adrenergic tone increased relative to the lower temperature. In conclusion, the cholinergic and adrenergic tones seem to be independent of temperature for colder thermal intervals (15-25°C), while exposure to a hotter ambient temperature (30°C) seems to be followed by a significant increase in adrenergic tone and may reflect cardiovascular adjustments made to match oxygen delivery to demand. Furthermore, while excluding the use of implantable electrodes or cannulae, this study provides a suitable non-invasive method for investigating cardiorespiratory function (cardiac and respiratory rates) in water-breathing animals such as the tadpole.

Hypoxia during embryonic development increases energy metabolism in normoxic juvenile chicks.

Environmental changes during perinatal development can affect the postnatal life. In this sense, chicken embryos that experience low levels of O2 over a specific phase of incubation can have their tissue growth reduced and the ventilatory response to hypoxia blunted, at least until hatching. Additionally, exposure to low level of O2 after birth reduces the thermogenesis as well. In the present study, we tested the hypothesis that hypoxia over the third week of incubation affects the thermoregulation of juvenile chicks at an age when thermogenesis is already expected to be well-developed. To this end, we measured body temperature (Tb) and oxygen consumption (V̇02) under acute hypoxia or different ambient temperatures (Ta) of 1 and 10day-old chicks that have been exposed to 21% O2 for entire incubation (Nx) or to 15% O2 in the last week of incubation (Hx). We also assessed the thermal preference under normoxia or acute hypoxia of the older chicks from both incubation groups in a thermocline. Hypoxia over incubation reduced growth but did not affect the cold-induced thermogenesis in hatchlings. Regarding the juvenile Hx, present data indicate a catch up growth with higher resting V̇02, a thermal preference for warmer Tas and a possible higher thermal conductance. In conclusion, our results show that hypoxia over the third week of incubation can affect the thermoregulation at least until 10days after hatch in chickens.

Acute effects of temperature and hypercarbia on cutaneous and branchial gas exchange in the South American lungfish, Lepidosiren paradoxa.

The South American lungfish, Lepidosiren paradoxa inhabits seasonal environments in the Central Amazon and Paraná-Paraguay basins that undergo significant oscillations in temperature throughout the year. They rely on different gas exchange organs, such as gills and skin for aquatic gas exchange while their truly bilateral lungs are responsible for aerial gas exchange; however, there are no data available on the individual contributions of the skin and the gills to total aquatic gas exchange in L. paradoxa. Thus, in the present study we quantify the relative contributions of skin and gills on total aquatic gas exchange during warm (35°C) and cold exposure (20°C) in addition to the effects of aerial and aquatic hypercarbia on aquatic gas exchange and gill ventilation rate (fG; 25°C), respectively. Elevated temperature (35°C) caused a significant increase in the contribution of cutaneous (from 0.61±0.13 to 1.34±0.26ml. STPD.h-1kg-1) and branchial (from 0.54±0.17 to 1.73±0.53ml. STPD.h-1kg-1) gas exchange for V̇CO2 relative to the lower temperature (20°C), while V̇O2 remained relatively unchanged. L. paradoxa exhibited a greater branchial contribution in relation to total aquatic gas exchange at lower temperatures (20 and 25°C) for oxygen uptake. Aerial hypercarbia decreased branchial V̇O2 whereas branchial V̇CO2 was significantly increased. Progressive increases in aquatic hypercarbia did not affect fG. This response is in contrast to increases in pulmonary ventilation that may offset any increase in arterial partial pressure of CO2 owing to CO2 loading through the animals' branchial surface. Thus, despite their reduced contribution to total gas exchange, cutaneous and branchial gas exchange in L. paradoxa can be significantly affected by temperature and aerial hypercarbia.

Orexinergic system in the locus coeruleus modulates the CO2 ventilatory response.

The orexins are hypothalamic neuropeptides involved in an array of functions such as regulation of sleep/wake states and chemoreception to CO2/pH. The locus coeruleus (LC) is a chemosensitive site and expresses an extensive population of orexin receptor 1 (OX1R). We tested the hypothesis that OX1Rs located in the LC participate in the ventilatory response to hypercapnia in a vigilance state and diurnal cycle-dependent manner. For this, we performed unilateral injections of SB-334867 (OX1R antagonist, 5 mM) into the LC of male Wistar rats and evaluated the ventilatory response to 7 % CO2 during wakefulness and sleep in the dark and light phases of the diurnal cycle. Hypercapnia induced an increase in ventilation (V E) in all groups compared to normocapnic values. However, during the dark phase, but not in the light phase, SB-334867 injection promoted an attenuation of the hypercapnic chemoreflex during wakefulness (V E: vehicle, 1502.6 ± 100 mL kg(-1) min(-1) vs SB-334867, 1200.3 ± 70.0 mL kg(-1) min(-1)) but not during sleep (V E: vehicle, 1383.0 ± 113.9 vs SB-334687, 1287.6 ± 92.1 mL kg(-1) min(-1)), due to changes in tidal volume (V T). We suggest that projections of orexin-containing neurons to the LC contribute, via OX1Rs, to the hypercapnic chemoreflex during wakefulness in the dark phase.

Nitric oxide and fever: immune-to-brain signaling vs. thermogenesis in chicks.

Nitric oxide (NO) plays a role in thermogenesis but does not mediate immune-to-brain febrigenic signaling in rats. There are suggestions of a different situation in birds, but the underlying evidence is not compelling. The present study was designed to clarify this matter in 5-day-old chicks challenged with a low or high dose of bacterial LPS. The lower LPS dose (2 µg/kg im) induced fever at 3-5 h postinjection, whereas 100 µg/kg im decreased core body temperature (Tc) (at 1 h) followed by fever (at 4 or 5 h). Plasma nitrate levels increased 4 h after LPS injection, but they were not correlated with the magnitude of fever. The NO synthase inhibitor (N(G)-nitro-l-arginine methyl ester, l-NAME; 50 mg/kg im) attenuated the fever induced by either dose of LPS and enhanced the magnitude of the Tc reduction induced by the high dose in chicks at 31-32°C. These effects were associated with suppression of metabolic rate, at least in the case of the high LPS dose. Conversely, the effects of l-NAME on Tc disappeared in chicks maintained at 35-36°C, suggesting that febrigenic signaling was essentially unaffected. Accordingly, the LPS-induced rise in the brain level of PGE2 was not affected by l-NAME. Moreover, l-NAME augmented LPS-induced huddling, which is indicative of compensatory mechanisms to run fever in the face of attenuated thermogenesis. Therefore, as in rats, systemic inhibition of NO synthesis attenuates LPS-induced fever in chicks by affecting thermoeffector activity and not by interfering with immune-to-brain signaling. This may constitute a conserved effect of NO in endotherms.

Baroreflex regulation affects ventilation in the cururu toad Rhinella schneideri.

Anurans regulate short-term oscillations in blood pressure through changes in heart rate (fH), vascular resistance and lymphatic fH Lung ventilation in anurans is linked to blood volume homeostasis by facilitating lymph return to the cardiovascular system. We hypothesized that the arterial baroreflex modulates pulmonary ventilation in the cururu toad Rhinella schneideri, and that this relationship is temperature dependent. Pharmacologically induced hypotension (sodium nitroprusside) and hypertension (phenylephrine) increased ventilation (25°C: 248.7±25.7 ml kg-1 min-1; 35°C: 351.5±50.2 ml kg-1 min-1) and decreased ventilation (25°C: 9.0±6.6 ml kg-1 min-1; 35°C: 50.7±15.6 ml kg-1 min-1), respectively, relative to control values from Ringer solution injection (25°C: 78.1±17.0 ml kg-1 min-1; 35°C: 137.7±15.5 ml kg-1 min-1). The sensitivity of the ventilatory response to blood pressure changes was higher during hypotension than during hypertension [25°C: -97.6±17.1 versus -23.6±6.0 breaths min-1 kPa-1; 35°C: -141.0±29.5 versus -28.7±6.4 breaths min-1 kPa-1, respectively; negative values indicate an inverse relationship between blood pressure and ventilation (or breathing frequency), i.e. as blood pressure increases, ventilation decreases, and vice versa], while temperature had no effect on these sensitivities. Hyperoxia (30%; 25°C) diminished ventilation, but did not abolish the ventilatory response to hypotension, indicating a response independent of peripheral chemoreceptors. Although there are previous data showing increased fH baroreflex sensitivity from 15 to 30°C in this species, further increases in temperature (35°C) diminished fH baroreflex gain (40.5±5.62 versus 21.6±4.64% kPa-1). Therefore, besides an involvement of pulmonary ventilation in matching O2 delivery to demand at higher temperatures in anurans, it also plays a role in blood pressure regulation, independent of temperature, possibly owing to an interaction between baroreflex and respiratory areas in the brain, as previously suggested for mammals.

Winter metabolic depression does not change arterial baroreflex control of heart rate in the tegu lizard Salvator merianae.

Baroreflex regulation of blood pressure is important for maintaining appropriate tissue perfusion. Although temperature affects heart rate (fH) reflex regulation in some reptiles and toads, no data are available on the influence of temperature-independent metabolic states on baroreflex. The South American tegu lizard Salvator merianae exhibits a clear seasonal cycle of activity decreasing fH along with winter metabolic downregulation, independent of body temperature. Through pharmacological interventions (phenylephrine and sodium nitroprusside), the baroreflex control of fH was studied at ∼ 25 °C in spring-summer- and winter-acclimated tegus. In winter lizards, resting and minimum fH were lower than in spring-summer animals (respectively, 13.3 ± 0.82 versus 10.3 ± 0.81 and 11.2 ± 0.65 versus 7.97 ± 0.88 beats min(-1)), while no acclimation differences occurred in resting blood pressure (5.14 ± 0.38 versus 5.06 ± 0.56 kPa), baroreflex gain (94.3 ± 10.7 versus 138.7 ± 30.3% kPa(-1)) or rate-pressure product (an index of myocardial activity). Vagal tone exceeded the sympathetic tone of fH, especially in the winter group. Therefore, despite the lower fH, winter acclimation does not diminish the fH baroreflex responses or rate-pressure product, possibly because of increased stroke volume that may arise because of heart hypertrophy. Independent of acclimation, fH responded more to hypotension than to hypertension. This should imply that tegus, which have no pressure separation within the single heart ventricle, must have other protection mechanisms against pulmonary hypertension or oedema, presumably through lymphatic drainage and/or vagal vasoconstriction of pulmonary artery. Such a predominant fH reflex response to hypotension, previously observed in anurans, crocodilians and mammals, may be a common feature of tetrapods.

Effect of temperature on chemosensitive locus coeruleus neurons of savannah monitor lizards, Varanus exanthematicus.

Savannah monitor lizards (Varanus exanthematicus) are unusual among ectothermic vertebrates in maintaining arterial pH nearly constant during changes in body temperature in contrast to the typical α-stat regulating strategy of most other ectotherms. Given the importance of pH in the control of ventilation, we examined the CO2/H+ sensitivity of neurons from the locus coeruleus (LC) region of monitor lizard brainstems. Whole-cell patch-clamp electrophysiology was used to record membrane voltage in LC neurons in brainstem slices. Artificial cerebral spinal fluid equilibrated with 80% O2, 0.0-10.0% CO2, balance N2, was superfused across brainstem slices. Changes in firing rate of LC neurons were calculated from action potential recordings to quantify the chemosensitive response to hypercapnic acidosis. Our results demonstrate that the LC brainstem region contains neurons that can be excited or inhibited by, and/or are not sensitive to CO2 in V. exanthematicus While few LC neurons were activated by hypercapnic acidosis (15%), a higher proportion of the LC neurons responded by decreasing their firing rate during exposure to high CO2 at 20°C (37%); this chemosensitive response was no longer exhibited when the temperature was increased to 30°C. Further, the proportion of chemosensitive LC neurons changed at 35°C with a reduction in CO2-inhibited (11%) neurons and an increase in CO2-activated (35%) neurons. Expressing a high proportion of inhibited neurons at low temperature may provide insights into mechanisms underlying the temperature-dependent pH-stat regulatory strategy of savannah monitor lizards.