Ampakine pretreatment enables a single brief hypoxic episode to evoke phrenic motor facilitation

Abstract

Phrenic long-term facilitation (LTF) is a sustained increase in phrenic motor output occurring after exposure to multiple (but not single) hypoxic episodes. Ampakines are a class of drugs that enhance AMPA receptor function. Ampakines can enhance expression of neuroplasticity, and the phrenic motor system is fundamentally dependent on excitatory glutamatergic currents. Accordingly, we tested the hypothesis that combining ampakine pretreatment with a single brief hypoxic exposure would result in phrenic motor facilitation lasting well beyond the period of hypoxia. Phrenic nerve output was recorded in urethane-anesthetized, ventilated, and vagotomized adult Sprague-Dawley rats. Ampakine CX717 (15 mg/kg iv; n = 8) produced a small increase in phrenic inspiratory burst amplitude and frequency, but values quickly returned to predrug baseline. When CX717 was followed 2 min later by a 5-min exposure to hypoxia (n = 8; ${\text{P}\text{a}}_{{\text{O}}_2}$ ~45 mmHg), a persistent increase in phrenic inspiratory burst amplitude (i.e., phrenic motor facilitation) was observed up to 60 min posthypoxia (103 ± 53% increase from baseline). In contrast, when hypoxia was preceded by vehicle injection (10% 2-hydroxypropyl-β-cyclodextrin; n = 8), inspiratory phrenic bursting was similar to baseline values at 60 min. Additional experiments with another ampakine (CX1739, 15 mg/kg) produced comparable results. We conclude that pairing low-dose ampakine treatment with a single brief hypoxic exposure can evoke sustained phrenic motor facilitation. This targeted approach for enhancing respiratory neuroplasticity may have value in the context of hypoxia-based neurorehabilitation strategies.NEW & NOTEWORTHY A single brief episode of hypoxia (e.g., 3-5 min) does not evoke long-lasting increases in respiratory motor output after the hypoxia is concluded. Ampakines are a class of drugs that enhance AMPA receptor function. We show that pairing low-dose ampakine treatment with a single brief hypoxic exposure can evoke sustained phrenic motor facilitation after the acute hypoxic episode.

Keywords: ampakine; neuroplasticity; respiratory.

Fig. 1.

Representative data from each experimental group. A: CX717 alone. B: CX717 followed by a single 5-min episode of hypoxia. C: vehicle injection 2-hydroxypropyl-β-cyclodextrin (HPCD) followed by a single 5-min episode of hypoxia. For each group, compressed records of instantaneous phrenic inspiratory burst frequency (top), arterial blood pressure (middle), and the phrenic neurogram (bottom) are shown at baseline, for 90 min following respective treatments, and during a single 5-min episode of hypoxia and maximum chemoreceptor activation (max challenge) at the end of each experiment (indicated by arrows).

Fig. 2.

Representative data illustrating the acute impact of ampakine delivery on phrenic motor output. Traces show that intravenous (I.V.) CX717 (A) caused an acute increase in phrenic nerve inspiratory burst amplitude. In contrast, vehicle injection (2-hydroxypropyl-β-cyclodextrin, HPCD; B) shows no apparent impact on phrenic output. Also depicted are arterial blood pressure and inspiratory burst rate.

Fig. 3.

Acute respiratory and cardiovascular effects of CX717 vs. vehicle (2-hydroxypropyl-β-cyclodextrin, HPCD). Peak effects of intravenous CX717or HPCD were measured 2 min after the start of the intravenous infusion. CX717 alone resulted in a significant increase in phrenic burst amplitude (CX717, P = 0.0467; HPCD, P = 0.4092; A), inspiratory burst frequency (CX717, P = 0.0010; HPCD, P = 0.5648; B), and mean arterial pressure (CX717, P = 0.0397; HPCD, P = 0.7729; D) but had no discernable impact on heart rate (CX717, P = 0.2214; HPCD, P = 0.4635; C). There was no acute impact of HPCD on any of these parameters. Individual data points are shown, and horizontal lines indicate means ± SD. +P < 0.05, statistically significant change from baseline conditions.

Fig. 4.

Pretreating with ampakine enables a single episode of hypoxia to trigger sustained facilitation of phrenic inspiratory burst amplitude. A: superimposed phrenic neurograms from each treatment group highlight the effect of CX717+ hypoxia (dark gray) on the amplitude of phrenic motor output compared with CX717alone (light gray) and 2-hydroxypropyl-β-cyclodextrin (HPCD) + hypoxia (black). Dotted line represents baseline amplitude. Arrows indicate points used for data analysis. B: average change in phrenic burst amplitude in the 3 experimental groups. Data from 90 min posttreatment, which were collected in a separate cohort, are shown in C. CX717 + hypoxia resulted in sustained increases in phrenic bursting that were 100% greater than baseline by 60 min. At the 90-min time point, average amplitude was significantly different in the CX717 + hypoxia group compared with the other groups and was significantly different from baseline. D: average change in burst amplitude from baseline during chemoreceptor challenge. Pretreatment with CX717 did not alter the amplitude of phrenic bursts during subsequent hypoxia (hypoxia 1) compared with the HPCD + hypoxia treatment group. After 60 min, the increase in burst amplitude during a second hypoxia challenge (hypoxia 2) as well as the maximal chemoreceptor challenge (max challenge) were elevated in the CX717+ hypoxia group. Individual data points are shown, and horizontal lines indicate means ± SD. #P < 0.05, significant difference between CX717 + hypoxia and HPCD + hypoxia. *P < 0.05, significant difference between CX717 and HPCD + hypoxia. ^P < 0.05, significant difference between CX717 and CX717 + hypoxia. +P < 0.05, significant difference compared with 0 (baseline).

Fig. 5.

Impact of ampakine CX717 on phrenic nerve burst frequency. A: average change in inspiratory phrenic burst frequency (Δbursts/min) from baseline after CX717 alone, CX717+ hypoxia, and 2-hydroxypropyl-β-cyclodextrin (HPCD + hypoxia). Data from 90 min posttreatment, which were collected in a separate cohort, are shown in B. Burst frequency was not significantly different at any time point in the HPCD + hypoxia group but was increased from baseline at 20 and 40 min after CX717 alone and at 40 and 60 min after CX717+ hypoxia. At 90 min, there was no significant difference in frequency between groups; however, in the CX717 + hypoxia group, frequency was significantly different from baseline. *P < 0.05, HPCD + hypoxia is significantly different from both other groups. +P < 0.05, significant difference compared with 0 (baseline).

Fig. 6.

Effects of ampakine CX717 on heart rate and mean arterial pressure. A: average change in heart rate (Δbeats/min) from baseline after CX717 alone, CX717+ hypoxia, and 2-hydroxypropyl-β-cyclodextrin (HPCD + hypoxia). Data from 90 min posttreatment, which were collected in a separate cohort, are shown in inset. There was no significant change in heart rate from baseline with CX717 alone; however, heart rate was lower than baseline at 40 and 60 min after CX717+ hypoxia and HPCD + hypoxia. At the 90-min time point, there were no differences between groups; however, the HPCD + hypoxia group was significantly different from baseline. B: average change in mean arterial pressure (ΔmmHg) from baseline. There was no significant decrease from baseline in mean arterial pressure in the CX717 group; however, average mean arterial pressure was decreased at 60 min after CX717 + hypoxia and at all time points after HPCD + hypoxia. At the 90-min time point, CX717 + hypoxia was significantly different from both other groups and significantly different from baseline. #P < 0.05, CX717 is significantly different from both other groups. *P < 0.05, HPCD + hypoxia is significantly different from both other groups. ^P < 0.05, CX717 + hypoxia is significantly different from both other groups. +P < 0.05, significant difference compared with 0 (baseline).

Fig. 7.

Results of supplemental experiments using ampakine CX1739. Phrenic nerve inspiratory burst amplitude was increased in rats receiving CX1739 + hypoxia vs. CX1739 alone (A). Inspiratory burst frequency also tended to be increased in the CX1739 + hypoxia group (B). Heart rate (C) and arterial blood pressure (D) were similar between groups. *P < 0.05, different from CX1739 alone.