A. G. Zabka

BDNF is necessary and sufficient for spinal respiratory plasticity following intermittent hypoxia

Intermittent hypoxia causes a form of serotonin-dependent synaptic plasticity in the spinal cord known as phrenic long-term facilitation (pLTF). Here we show that increased synthesis of brain-derived neurotrophic factor (BDNF) in the spinal cord is necessary and sufficient for pLTF in adult rats. We found that intermittent hypoxia elicited serotonin-dependent increases in BDNF synthesis in ventral spinal segments containing the phrenic nucleus, and the magnitude of these BDNF increases correlated with pLTF magnitude. We used RNA interference (RNAi) to interfere with BDNF expression, and tyrosine kinase receptor inhibition to block BDNF signaling. These disruptions blocked pLTF, whereas intrathecal injection of BDNF elicited an effect similar to pLTF. Our findings demonstrate new roles and regulatory mechanisms for BDNF in the spinal cord and suggest new therapeutic strategies for treating breathing disorders such as respiratory insufficiency after spinal injury. These experiments also illustrate the potential use of RNAi to investigate functional consequences of gene expression in the mammalian nervous system in vivo.

Sex steroid hormones and the neural control of breathing

We review evidence that sex steroid hormones including estrogen, progesterone and testosterone are involved in the central neural control of breathing. Sex hormones may exert their effects on respiratory motoneurons via neuromodulators, in particular, the serotonergic system. Recent studies have shown that levels of serotonin (5HT) in the hypoglossal and phrenic nuclei are greater in female than in male rats. Serotonin-dependent plasticity in hypoglossal and phrenic motor output also differs in male and female rats. Changing levels of gonadal hormones throughout the estrus cycle coincide with changing levels of 5HT in respiratory motor nuclei, and gonadectomy in male rats results in a decrease in 5HT-dependent plasticity in respiratory motor output. We speculate that sex steroid hormones are critically involved in adaptations in the neural control of breathing throughout life, and that decreasing levels of these hormones with increasing age may have a negative influence on the respiratory control system in response to challenge.

Long term facilitation of respiratory motor output decreases with age in male rats

1 Long term facilitation (LTF) is a serotonin‐dependent augmentation of respiratory motor output (phrenic and hypoglossal) following episodic hypoxia. Since ageing influences respiratory control mechanisms and serotonergic function, we tested the hypothesis that LTF decreases with age in male rats. 2 Young (3‐4 month) and aged (13 month) male Sprague‐Dawley rats were anaesthetized with urethane, vagotomized, paralysed and pump ventilated. Integrated phrenic and hypoglossal (XII) nerve activities were measured before (baseline), during and for 60 min after three 5 min episodes of isocapnic hypoxia (Pa,O2 35‐45 mmHg) separated by 5 min of hyperoxia (Pa,O2 > 150 mmHg). 3 In young rats, LTF was observed as an augmentation in peak integrated phrenic (n = 8) and XII (n = 7) amplitudes following episodic hypoxia (56 ± 14 and 73 ± 16 % (means ±s.e.m.) at 60 min post‐hypoxia, respectively; both P < 0.05). In aged rats, LTF was significantly increased compared to baseline in phrenic (25 ± 8 % at 60 min, P < 0.05), but not in XII (4 ± 7 %, P > 0.05) motor output. LTF was significantly greater in young than in aged rats in both motor outputs (P < 0.05). 4 Decreased phrenic and XII LTF suggests that serotonergic modulation of respiratory motor output decreases in ageing male rats. We speculate that decreased serotonergic modulation may contribute to age‐related breathing disorders.

Respiratory plasticity: differential actions of continuous and episodic hypoxia and hypercapnia.

The objectives of this paper are: (1) to review advances in our understanding of the mechanisms of respiratory plasticity elicited by episodic versus continuous hypoxia in short to intermediate time domains (min to h); and (2) to present new data suggesting that different patterns of hypercapnia also elicit distinct forms of respiratory plasticity. Episodic, but not continuous hypoxia elicits long-term facilitation (LTF) of respiratory motor output. Phrenic LTF is a serotonin-dependent central neural mechanism that requires: (a) activation of spinal serotonin receptors; and (b) spinal protein synthesis. Continuous and episodic hypercapnia also elicit different mechanisms of plasticity. Continuous, severe hypercapnia (25 min of approximately 10% inspired CO(2)) elicits long-term depression (LTD) of phrenic motor output (-33+/-8% at 60 min post-hypercapnia) in anesthetized rats. In contrast, 3,5 min hypercapnic episodes do not elicit LTD (9+/-17% at 60 min). We hypothesize that the response of respiratory motoneurons to serotonergic and noradrenergic modulation may contribute to pattern sensitivity to hypoxia and hypercapnia.

Age and gender effects on serotonin-dependent plasticity in respiratory motor control

We review recent evidence indicating that serotonin-dependent plasticity in respiratory control is influenced by age and gender. Specifically, respiratory long-term-facilitation following intermittent hypoxia decreases with age in male rats, but increases in female rats. We speculate about a possible relationship between age and gender effects on serotonin-dependent plasticity in upper airway motoneuron pools and the prevalence of obstructive sleep apnea.

Ageing and gonadectomy have similar effects on hypoglossal long‐term facilitation in male Fischer rats

Long‐term facilitation (LTF), a form of serotonin‐dependent respiratory plasticity induced by intermittent hypoxia, decreases with increasing age or following gonadectomy in male Sprague‐Dawley (SD) rats. Ageing is accompanied by decreasing levels of testosterone, which in turn influences serotonergic function. In addition, LTF in young male rats differs among strains. Thus, we tested the hypothesis that LTF is similar in middle‐aged and gonadectomized young male rats of an inbred rat strain commonly used in studies on ageing (F344) by comparison with SD rats. We further tested whether the magnitude of LTF correlates with circulating serum levels of testosterone and/or progesterone. Young and middle‐aged intact and young gonadectomized (GDX) male Fischer 344 rats were anaesthetized, neuromuscularly blocked and ventilated. Integrated phrenic and hypoglossal (XII) nerve activities were measured before, during and 60 min following three 5‐min episodes of isocapnic hypoxia. LTF was observed in phrenic motor output in young and middle‐aged intact and young GDX rats. In contrast, XII LTF was observed only in young intact rats. In middle‐aged and young GDX rats, XII LTF was significantly lower than in young intact rats (P < 0.05). Furthermore, XII LTF was positively correlated with the testosterone/progesterone ratio. These data show that serotonin‐dependent plasticity in upper airway respiratory output is similar in F344 and SD rat strains. Furthermore, LTF is similarly impaired in middle‐aged and gonadectomized male rats, suggesting that gonadal hormones play an important role in modulating the capacity for neuroplasticity in upper airway motor control.

Conversion from testosterone to oestradiol is required to modulate respiratory long‐term facilitation in male rats

Sex hormones modulate plasticity in the central nervous system, including respiratory long‐term facilitation (LTF), a form of serotonin‐dependent respiratory plasticity induced by intermittent hypoxia. Since gonadectomy (GDX) attenuates LTF in male rats, we tested the hypotheses that: (1) testosterone replenishment restores LTF in gonadectomized male rats, and (2) that the conversion of testosterone to oestradiol (under the influence of aromatase) is required for these effects. Intact and sham operated male F344 rats were compared to gonadectomized rats implanted with Silastic tubing containing testosterone (T), T plus an aromatase inhibitor (ADT), or 5α‐dihydrotestosterone (DHT), a form of testosterone not converted to oestradiol. Seven days postsurgery, LTF was studied in anaesthetized, neuromuscularly blocked and ventilated rats while monitoring integrated phrenic and hypoglossal (XII) motor output. LTF was elicited by three 5 min hypoxic episodes (P  a,O 2 = 35 − 45 mmHg). Although significant phrenic and XII LTF were observed in all rat groups, GDX reduced both phrenic and XII LTF, an effect reversed by T. In contrast, LTF was not restored in T + ADT or DHT‐treated gonadectomized rats. We conclude that the conversion of testosterone to oestradiol modulates phrenic and XII LTF in male F344 rats.

Age and gender effects on serotonergic innervation and modulation of the hypoglossal motor nucleus

Aging results in structural, functional and neurochemical alterations in the respiratory system. Serotonin (5HT) plays a major role in breathing and the control of upper airway function. We tested the hypothesis that with increasing age there is a selective decrease in serotonergic modulation of respiratory motoneurons, in particular hypoglossal motoneurons to the tongue in male rats. We used light microscopic immunocytochemistry to study the distribution of 5HT axons and boutons throughout the hypoglossal nucleus in young and old rats male and female rats. Aged male rats (>12 months) had fewer serotonin immunoreactive axons and boutons in the hypoglossal nucleus than young male rats (<6 months). In contrast, 5HT immunoreactivity in the hypoglossal nucleus in female rats was higher than in age-matched males, and increased with age. In order to assess the functional consequences of this anatomical reorganization, we measured long term facilitation (LTF), a serotonin-dependent, long lasting increase in respiratory motor output following episodic hypoxia. LTF in the hypoglossal motor output was significantly reduced in aged male rats by comparison with young male rats [1]. In contrast, hypoglossal LTF increased in aged female rats in diestrus, but not in estrus. Taken together, these data suggest that in males, but not females, normal aging may result in decreased serotonergic facilitation of hypoglossal motoneurons that could result in reduced airway patency.