Concentration-related metabolic rate and behavioral thermoregulatory adaptations to serial administrations of nitrous oxide in rats

Abstract

Background: Initial administration of ≥60% nitrous oxide (N2O) to rats evokes hypothermia, but after repeated administrations the gas instead evokes hyperthermia. This sign reversal is driven mainly by increased heat production. To determine whether rats will behaviorally oppose or assist the development of hyperthermia, we previously performed thermal gradient testing. Inhalation of N2O at ≥60% causes rats to select cooler ambient temperatures both during initial administrations and during subsequent administrations in which the hyperthermic state exists. Thus, an available behavioral response opposes (but does not completely prevent) the acquired hyperthermia that develops over repeated high-concentration N2O administrations. However, recreational and clinical uses of N2O span a wide range of concentrations. Therefore, we sought to determine the thermoregulatory adaptations to chronic N2O administration over a wide range of concentrations.

Methods: This study had two phases. In the first phase we adapted rats to twelve 3-h N2O administrations at either 0%, 15%, 30%, 45%, 60% or 75% N2O (n = 12 per group); outcomes were core temperature (via telemetry) and heat production (via respirometry). In the second phase, we used a thermal gradient (range 8°C-38°C) to assess each adapted group's thermal preference, core temperature and locomotion on a single occasion during N2O inhalation at the assigned concentration.

Results: In phase 1, repeated N2O administrations led to dose related hyperthermic and hypermetabolic states during inhalation of ≥45% N2O compared to controls (≥ 30% N2O compared to baseline). In phase 2, rats in these groups selected cooler ambient temperatures during N2O inhalation but still developed some hyperthermia. However, a concentration-related increase of locomotion was evident in the gradient, and theoretical calculations and regression analyses both suggest that locomotion contributed to the residual hyperthermia.

Conclusions: Acquired N2O hyperthermia in rats is remarkably robust, and occurs even despite the availability of ambient temperatures that might fully counter the hyperthermia. Increased locomotion in the gradient may contribute to hyperthermia. Our data are consistent with an allostatic dis-coordination of autonomic and behavioral thermoregulatory mechanisms during drug administration. Our results have implications for research on N2O abuse as well as research on the role of allostasis in drug addiction.

Fig 1. Temporal profiles depicting concentration-dependent thermal adaptations to 12 steady-state nitrous oxide exposures in rats.

The time plots depict mean core temperature and body-mass adjusted heat production in the first and twelfth 3-h N₂O exposure sessions (indicated by the numbers in each concentration group). Bars indicate period of N₂O inhalation. Thin lines depict outcomes during control gas (custom control air, o% N₂O) administration; dashed thin line denotes first control session, solid thin line, denotes twelfth control session). N = 12 per dose group. During the course of the calorimetry studies, body mass increased by ~125% whereas baseline HP increased by ~35% (error bars are 95% confidence intervals). The regression equation for HP was used to adjust HP for body mass in the time series plots (see Materials and Methods). N₂O, nitrous oxide; Tcore, core temperature; HP, heat production.

Fig 2

Nitrous oxide minus control gas differences in core temperature and heat production in each of the twelve 3-h exposure sessions (first 90 min). 95% confidence intervals that do not include zero are significantly different from control at p<0.05. N = 12 per dose group. C.I., confidence interval; N₂O, nitrous oxide; Tcore, core temperature; HP, heat production.

Fig 3. Nitrous oxide minus control gas differences in core temperature and heat production in each of the twelve 3-h exposure sessions (second 90 min).

For explanation see legend for Fig 2.

Fig 4. Time series plots of core temperature and selected ambient temperature during a single nitrous oxide exposure in the thermally graded alleyway following 12 drug administrations in the calorimetry phase of the study.

Despite access to ambient temperatures that presumably would have prevented hyperthermia (8°C at the coldest end), rats in the higher dose groups selected only modestly lower temperatures and still developed hyperthermia in the earlier part of administration. N = 12 per group. Data are mean ± 95% confidence limits. Tcore, core temperature; Tsel, selected ambient temperature; N₂O, nitrous oxide.

Fig 5. Analysis of baseline-adjusted mean changes in core temperature and selected ambient temperature during nitrous oxide exposure in the thermally graded alleyway.

95% confidence intervals that do not include zero are significantly different from baseline at p<0.05. Numbers above error flags are p-values for the group differences from the control gas condition. N = 12 per group. Overall baseline core temperature was 37.0°C (95% C.I. = 36.9, 37.1) while baseline selected temperature was 24.9°C (95% C.I. = 24.3, 25.7). Tcore, core temperature; Tsel, selected ambient temperature; N₂O, nitrous oxide; C.I. confidence interval.

Fig 6. Concentration-related stimulatory effect of nitrous oxide on total locomotion and its estimated influence on core temperature during thermal gradient testing.

Data are medians bounded by 25^th and 75^th percentiles. P-values for comparisons with control gas condition are indicated. Estimated changes in core temperature are based only on minimum cost of transport and ignore the influence of resting metabolic rate and the cost of support (see Materials and Methods); hence the total contribution of locomotion to body heat content is probably higher. N = 12 per group. Tcore, core temperature.