Adenosine and γ-aminobutyric acid partially regulate metabolic and ventilatory responses of Damaraland mole-rats to acute hypoxia.

Fossorial Damaraland mole-rats (Fukomys damarensis) mount a robust hypoxic metabolic response (HMR) but a blunted hypoxic ventilatory response (HVR) to acute hypoxia. Although these reflex physiological responses have been described previously, the underlying signalling pathways are entirely unknown. Of particular interest are contributions from γ-aminobutyric acid (GABA), which is the primary inhibitory neurotransmitter in the nervous system of most adult mammals, and adenosine, the accumulation of which increases during hypoxia as a breakdown product of ATP. Therefore, we hypothesized that GABAergic and/or adenosinergic signalling contributes to the blunted HVR and robust HMR in Damaraland mole-rats. To test this hypothesis, we injected adult animals with saline alone (controls), or 100 mg kg-1 aminophylline or 1 mg kg-1 bicuculline, to block adenosine or GABAA receptors, respectively. We then used respirometry, plethysmography and thermal RFID probes to non-invasively measure metabolic, ventilator and thermoregulatory responses, respectively, to acute hypoxia (1 h in 5 or 7% O2) in awake and freely behaving animals. We found that bicuculline had relatively minor effects on metabolism and thermoregulation but sensitized ventilation such that the HVR became manifest at 7% instead of 5% O2 and was greater in magnitude. Aminophylline increased metabolic rate, ventilation and body temperature in normoxia, and augmented the HMR and HVR. Taken together, these findings indicate that adenosinergic and GABAergic signalling play important roles in mediating the robust HMR and blunted HVR in Damaraland mole-rats.

The glutamatergic drive to breathe is reduced in severe but not moderate hypoxia in Damaraland mole-rats.

Damaraland mole-rats (Fukomys damarensis) are a hypoxia-tolerant fossorial species that exhibit a robust hypoxic metabolic response (HMR) and blunted hypoxic ventilatory response (HVR). Whereas the HVR of most adult mammals is mediated by increased excitatory glutamatergic signalling, naked mole-rats, which are closely related to Damaraland mole-rats, do not utilize this pathway. Given their phylogenetic relationship and similar lifestyles, we hypothesized that the signalling mechanisms underlying physiological responses to acute hypoxia in Damaraland mole-rats are like those of naked mole-rats. To test this, we used pharmacological antagonists of glutamatergic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) and N-methyl-d-aspartate receptors (NMDARs), combined with plethysmography, respirometry and thermal RFID chips, to non-invasively evaluate the role of excitatory AMPAR and NMDAR signalling in mediating ventilatory, metabolic and thermoregulatory responses, respectively, to 1 h of 5 or 7% O2. We found that AMPAR or NMDAR antagonism have minimal impacts on the HMR or hypoxia-mediated changes in thermoregulation. Conversely, the 'blunted' HVR of Damaraland mole-rats is reduced by either AMPAR or NMDAR antagonism such that the onset of the HVR occurs in less severe hypoxia. In more severe hypoxia, antagonists have no impact, suggesting that these receptors are already inhibited. Together, these findings indicate that the glutamatergic drive to breathe decreases in Damaraland mole-rats exposed to severe hypoxia. These findings differ from other adult mammals, in which the glutamatergic drive to breathe increases with hypoxia.

Physiological responses to hypoxia are constrained by environmental temperature in heterothermic tenrecs.

Malagasy tenrecs are placental hibernating mammals that seal the entrances to their burrows and hibernate either singly or in groups for 8-9 months, which is likely to create a hypoxic and hypercapnic burrow environment. Therefore, we hypothesized that tenrecs are tolerant to environmental hypoxia and hypercapnia. Many hypoxia- and hypercapnia-tolerant fossorial mammals respond to hypoxia by decreasing metabolic rate and thermogenesis, and have blunted ventilatory responses to both environmental hypoxia and hypercapnia. However, tenrecs exhibit extreme metabolic and thermoregulatory plasticity, which exceeds that of most heterothermic mammals and approaches that of ectothermic reptiles. Thus, we predicted that tenrecs would have abnormal physiological responses to hypoxia and hypercapnia relative to other fossorial mammals. To test this, we exposed common tenrecs (Tenrec ecaudatus) to moderate and severe hypoxia (9 and 4% O2) or hypercapnia (5 and 10% CO2) in either 28 or 16°C while non-invasively measuring metabolic rate, thermogenesis and ventilation. We found that tenrecs exhibit robust metabolic decreases in both hypoxia and hypercapnia. Furthermore, tenrecs have blunted ventilatory responses to both hypoxia and hypercapnia, and these responses are highly temperature sensitive such that they are reduced or absent in 16°C. Thermoregulation was highly variable in 16°C but constrained in 28°C across all treatment conditions and was not impacted by hypoxia or hypercapnia, unlike in other heterothermic mammals. Taken together, our results indicate that physiological responses to hypoxia and hypercapnia in tenrecs are highly dependent on environmental temperature and differ from those of other mammalian heterotherms.

Burrowing star-nosed moles (Condylura cristata) are not hypoxia tolerant.

Star-nosed moles (Condylura cristata) have an impressive diving performance and burrowing lifestyle, yet no ventilatory data are available for this or any other talpid mole species. We predicted that, like many other semi-aquatic and fossorial small mammals, star-nosed moles would exhibit: (i) a blunted (i.e. delayed or reduced) hypoxic ventilatory response, (ii) a reduced metabolic rate and (iii) a lowered body temperature (Tb) in hypoxia. We thus non-invasively measured these variables from wild-caught star-nosed moles exposed to normoxia (21% O2) or acute graded hypoxia (21-6% O2). Surprisingly, star-nosed moles did not exhibit a blunted HVR or decreased Tb in hypoxia, and only manifested a significant, albeit small (<8%), depression of metabolic rate at 6% O2 relative to normoxic controls. Unlike small rodents inhabiting similar niches, star-nosed moles are thus intolerant to hypoxia, which may reflect an evolutionary trade-off favouring the extreme sensory biology of this unusual insectivore.

Na+/K+-ATPase activity is regionally regulated by acute hypoxia in naked mole-rat brain.

Matching ATP supply and demand is key to neuronal hypoxia-tolerance and failure to achieve this balance leads to excitotoxic cell death in most adult mammalian brains. Ion pumping is the most energy-demanding process in the brain and some hypoxia-tolerant vertebrates coordinately down-regulate ion movement across neuronal membranes to reduce the workload of energy-expensive ion pumps, and particularly the Na+/K+-ATPase. Naked mole-rats are among the most hypoxia-tolerant mammals and achieve a hypometabolic state while maintaining brain [ATP] during severe hypoxia; however, whether ionic homeostasis is plastic in naked mole-rat brain is unknown. To examine this question, we exposed animals to 4 h of normoxia or moderate or severe hypoxia (11 or 3% O2, respectively) and measured changes in brain Na+/K+-ATPase activity. We found that 1) whole body metabolic rate decreased ∼25 and 75% in moderate and severe hypoxia, respectively, and 2) Na+/K+-ATPase activity decreased ∼50% in forebrain but increased 2-fold in cerebellum and was unchanged in brainstem. These results indicate that naked mole-rats acutely modulate brain energy demand in a region-specific manner to prioritize energy usage by the cerebellum. This may support exploration, navigation, and escape behaviours, while also enabling ATP savings when encountering hypoxia in nature.

Naked mole-rats suppress energy metabolism and modulate membrane cholesterol in chronic hypoxia.

Naked mole-rats (NMRs) are mammalian champions of hypoxia tolerance that enter metabolic suppression to survive in low oxygen environments. Common physiological mechanisms used by animals to suppress metabolic rate include downregulating energy metabolism (ATP supply) as well as ion pumps (primary cellular ATP consumers). A recent goldfish study demonstrated that remodeling of membrane lipids may mediate these responses, but it is unknown if NMR employs the same strategies; therefore, we aimed to test the hypotheses that these fossorial mammals 1) downregulate the activity of key enzymes of glycolysis, tricarboxylic acid (TCA) cycle, and ß-oxidation, 2) inhibit sodium-potassium-ATPase, and 3) alter membrane lipids in response to chronic hypoxia. We found that NMRs exposed to 11% oxygen for 4 wk had a lower metabolic rate by 34%. This suppression occurs concurrently with tissue-specific 25-99% decreases in metabolic enzymes activities, a 77% decrease in brain sodium/potassium-ATPase activity, and widespread changes in membrane cholesterol abundance. By reducing glycolytic and ß-oxidation fluxes, NMRs decrease the supply of acetyl-CoA to the TCA cycle. By contrast, there is a 94% upregulation of citrate synthase in the heart, possibly to support circulation and thus oxygen supply to other organs. Taken together, these responses may reflect a coordinated physiological response to hypoxia, but a clear functional link between changes in membrane composition and enzyme activities could not be established. Nevertheless, this is the first demonstration that hypometabolic NMRs alter the lipid composition of their membranes in response to chronic in vivo exposure to hypoxia.

Fossorial giant Zambian mole-rats have blunted ventilatory responses to environmental hypoxia and hypercapnia.

Fossorial giant Zambian mole-rats are believed to live in a hypoxic and hypercapnic subterranean environment but their physiological responses to these challenges are entirely unknown. To investigate this, we exposed awake and freely-behaving animals to i) 6 h of normoxia, ii) acute graded normocapnic hypoxia (21, 18, 15, 12, 8, and 5% O2, 0% CO2, balance N2; 1 h each), or iii) acute graded normoxic hypercapnia (0, 2, 5, 7, 9, and 10% CO2, 21% O2, balance N2; 1 h each), followed by a 1 h normoxic normocapnic recovery period, while non-invasively measuring ventilation, metabolic rate, and body temperature (Tb). We found that these mole-rats had a blunted hypoxic ventilatory response that manifested at 12% inhaled O2, a robust hypoxic metabolic response (up to a 68% decrease, starting at 15% O2), and decreased Tb (at or below 8% O2). Upon reoxygenation, metabolic rate increased 52% above normoxic levels, suggesting the paying off of an O2 debt. Ventilation was less sensitive to environmental hypercapnia than to environmental hypoxia and animals also exhibited a blunted hypercapnic ventilatory response that did not manifest below 9% inhaled CO2. Conversely, metabolism and Tb were not affected by hypercapnia. Taken together, these results indicate that, like most other fossorial rodents, giant Zambian mole-rats have blunted hypoxic and hypercapnic ventilatory responses and employ metabolic suppression to tolerate acute hypoxia. Blunted physiological responses to hypoxia and hypercapnia likely reflect the subterranean lifestyle of this mammal, wherein intermittent but severe hypoxia and/or hypercapnia may be common challenges.

Neurokinin-1 receptor activation is sufficient to restore the hypercapnic ventilatory response in the Substance P-deficient naked mole-rat.

Naked mole-rats (NMRs) live in large colonies within densely populated underground burrows. Their collective respiration generates significant metabolic carbon dioxide (CO2) that diffuses slowly out of the burrow network, creating a hypercapnic environment. Currently, the physiological mechanisms that underlie the ability of NMRs to tolerate environmental hypercapnia are largely unknown. To address this, we used whole-body plethysmography and respirometry to elucidate the hypercapnic ventilatory and metabolic responses of awake, freely behaving NMRs to 0%-10% CO2. We found that NMRs have a blunted hypercapnic ventilatory response (HCVR): ventilation increased only in 10% CO2. Conversely, metabolism was unaffected by hypercapnia. NMRs are insensitive to cutaneous acid-based pain caused by modified substance P (SP)-mediated peripheral neurotransmission, and SP is also an important neuromodulator of ventilation. Therefore, we re-evaluated physiological responses to hypercapnia in NMRs after an intraperitoneal injection of exogenous substance P (2 mg/kg) or a long-lived isoform of substance P {[pGlu5-MePhe8-MeGly9]SP(5-11), DiMe-C7; 40-400 µg/kg}. We found that both drugs restored hypercapnia sensitivity and unmasked an HCVR in animals breathing 2%-10% CO2. Taken together, our findings indicate that NMRs are remarkably tolerant of hypercapnic environments and have a blunted HCVR; however, the signaling network architecture required for a "normal" HCVR is retained but endogenously inactive. This muting of chemosensitivity likely suits the ecophysiology of this species, which presumably experiences hypercapnia regularly in their underground niche.