Sex hormone supplementation improves breathing and restores respiratory neuroplasticity following C2 hemisection in rats.

In addition to loss of sensory and motor function below the level of the lesion, traumatic spinal cord injury (SCI) may reduce circulating steroid hormones that are necessary for maintaining normal physiological function for extended time periods. For men, who comprise nearly 80% of new SCI cases each year, testosterone is the most abundant circulating sex steroid. SCI often results in significantly reduced testosterone production and may result in chronic low testosterone levels. Testosterone plays a role in respiratory function and the expression of respiratory neuroplasticity. When testosterone levels are low, young adult male rats are unable to express phrenic long-term facilitation (pLTF), an inducible form of respiratory neuroplasticity invoked by acute, intermittent hypoxia (AIH). However, testosterone replacement can restore this respiratory neuroplasticity. Complicating the interpretation of this finding is that testosterone may exert its influence in three possible ways: 1) directly through androgen receptor (AR) activation, 2) through conversion to dihydrotestosterone (DHT) by way of the enzyme 5α-reductase, or 3) through conversion to 17ß-estradiol (E2) by way of the enzyme aromatase. DHT signals via AR activation similar to testosterone, but with higher affinity, while E2 activates local estrogen receptors. Evidence to date supports the idea that exogenous testosterone supplementation exerts its influence through estrogen receptor signaling under conditions of low circulating testosterone. Here we explored both recovery of breathing function (measured with whole body barometric plethysmography) and the expression of AIH-induced pLTF in male rats following C2-hemisection SCI. One week post injury, rats were supplemented with either E2 or DHT for 7 days. We hypothesized that E2 would enhance ventilation and reveal pLTF following AIH in SCI rats. To our surprise, though E2 did beneficially impact overall breathing recovery following C2-hemisection, both E2 supplementation and DHT restored the expression of AIH-induced pLTF 2 weeks post-SCI.

Low level CO2 supplementation maintains isocapnia and reveals ventilatory long-term facilitation in rats.

Acute, intermittent hypoxia (AIH) induces ventilatory long-term facilitation (vLTF) in awake, freely behaving rats under poikilocapnic and isocapnic experimental conditions. Establishing pre-clinical methods for vLTF induction that more closely align with successful protocols in humans and anesthetized rats would minimize dissonance in experimental findings and improve translational aspects of vLTF. Here, we tested several levels of low-dose CO2 supplementation during and after AIH to determine 1) the lowest amount of inspired CO2 that would maintain isocapnia in rats during a vLTF protocol, and 2) the net impact of supplemental CO2 on vLTF expression. Rats received one of four levels of inspired CO2 (0%, 0.5%, 1% or 2%) administered during AIH and for the 60 min following AIH to quantify vLTF. Our findings indicated that 2% inspired CO2 was sufficient to maintain isocapnia across the AIH protocol and reveal significant vLTF. These findings provide evidence-based support for using 2% supplemental CO2 during and after AIH when assessing vLTF in rats.

The long-term impact of ovariectomy on ventilation and expression of phrenic long-term facilitation in female rats.

NEW FINDINGS: What is the central question of this study? Would ovariectomy cause prolonged changes in ventilation and sustained loss of acute, intermittent hypoxia-induced neuroplasticity or would these outcomes be restored with time? What is the main finding and its importance? Our main findings demonstrate that ovariectomy elicits minimal alteration in overall breathing function but impairs acute, intermittent hypoxia-induced plasticity for ≤ 12 weeks. ABSTRACT: Sex hormones are necessary to enable respiratory neuroplasticity, including phrenic long-term facilitation (pLTF), a form of respiratory motor plasticity elicited by acute, intermittent hypoxia (AIH). Female rats exhibit a progressive increase in phrenic nerve amplitude after AIH characteristic of pLTF only during pro-oestrus, the stage of the oestrous cycle notable for elevated circulating oestradiol levels. Removal of the ovaries [ovariectomy (OVX)], the primary source of circulating oestradiol, also eliminates AIH-induced pLTF after 1 week. Ovariectomy is used routinely as a model to examine the impact of sex hormones on CNS structure and function, but the long-term impact of OVX is rarely examined. Extra-ovarian sites of oestradiol synthesis, including multiple CNS sites, have been identified and might possess the capacity to restore oestradiol levels, in part, over time, impacting respiratory function and the expression of respiratory neuroplasticity. We examined both ventilation in awake, freely behaving female rats, using barometric plethysmography, and the expression of AIH-induced pLTF in anaesthetized, ventilated female rats 2 and 12 weeks after OVX and compared them with age-matched ovarian-intact female rats. Our findings indicate that chronic OVX had little impact on baseline breathing or in the response to respiratory challenge (10% O2 , 5% CO2 , balance N2 ) during plethysmography. However, OVX rats at both 2 and 12 weeks demonstrated a persistent loss of AIH-induced pLTF relative to control animals (P < 0.01), suggesting that other sources of oestradiol synthesis were insufficient to restore pLTF. These data are consistent with our previous work indicating that oestradiol plays a key role in expression of AIH-induced respiratory neuroplasticity.