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Tauopathy comprises hyperphosphorylation of the microtubule-associated protein tau, causing intracellular aggregation and accumulation as neurofibrillary tangles and neuropil treads. Some primary tauopathies are linked to mutations in the MAPT gene coding for protein tau, but most are sporadic with unknown causes. Also, in Alzheimer's disease, the most frequent secondary tauopathy, neither the cause nor the pathological mechanisms and repercussions are understood. Transgenic mice expressing mutant Tau-P301L suffer cognitive and motor defects and die prematurely from unknown causes. Here, in situ electrophysiology in symptomatic Tau-P301L mice (7-8 months of age) revealed reduced postinspiratory discharges of laryngeal motor outputs that control laryngeal constrictor muscles. Under high chemical drive (hypercapnia), postinspiratory discharge was nearly abolished, whereas laryngeal inspiratory discharge was increased disproportionally. The latter may suggest a shift of postinspiratory laryngeal constrictor activity into inspiration. In vivo double-chamber plethysmography of Tau-P301L mice showed significantly reduced respiratory airflow but significantly increased chest movements during baseline breathing, but particularly in hypercapnia, confirming a significant increase in inspiratory resistive load. Histological analysis demonstrated hyperphosphorylated tau in brainstem nuclei, directly or indirectly involved in upper airway motor control (i.e., the Kölliker-Fuse, periaqueductal gray, and intermediate reticular nuclei). In contrast, young Tau-P301L mice did not show breathing disorders or brainstem tauopathy. Consequently, in aging Tau-P301L mice, progressive upper airway dysfunction is linked to progressive tauopathy in identified neural circuits. Because patients with tauopathy suffer from upper airway dysfunction, the Tau-P301L mice can serve as an experimental model to study disease-specific synaptic dysfunction in well defined functional neural circuits.

Experiments were carried out on the in vitro brainstem-spinal cord preparation of the newborn rat to analyse the effects of substance P (SP) on phrenic motoneuron (PMN) activity. In current-clamp mode, SP significantly depolarized PMNs, increased their input resistance, decreased the rheobase current and shifted the firing frequency-intensity relationships leftwards, but did not affect spike frequency adaptation or single spike configuration. The neurokinin receptor agonist NK1 had SP-mimetic effects, whereas the NK3 and NK2 receptor agonists were less effective and ineffective, respectively. In a tetrodotoxin-containing aCSF, only SP or the NK1 receptor agonist were still active. No depolarization was observed when the NK1 receptor agonist was applied in the presence of muscarine. In voltage-clamp mode, SP or the NK1 receptor agonist produced an inward current (ISP) which was not significantly reduced by extracellular application of tetraethylammonium, Co2+, 4-aminopyridine or Cs+. In aCSF containing tetrodotoxin, Co2+ and Cs+, ISP was blocked by muscarine. No PMN displayed any M-type potassium current but only a current showing no voltage sensitivity over the range -100 to 0 mV, reversing near the expected EK +, hence consistent with a leak current. SP application to the spinal cord only (using a partitioned chamber) significantly increased the phrenic activity. Pretreatment with the NMDA receptor antagonist DL-2-amino-5-phosphonovaleric acid (AP5) decreased the C4 discharge duration and blocked the effect of SP, thus exhibiting an NMDA potentiation by SP. In conclusion, SP modulates postsynaptically the response of phrenic motoneurons to the inspiratory drive through the reduction of a leak conductance and the potentiation of the NMDA component of the synaptic input.

Experiments were performed on neonatal mice to know whether substance P (SP) modified the rhythm and the amplitude of the phrenic bursts generated in vitro in brainstem-cervical cord preparations. In OF1 and C3H neonatal preparations, SP or the tachykinin NK1 receptor agonist [Sar9,Met(O2)11] substance P both increased significantly phrenic burst amplitude (10(-7) M) but had no significant effect on respiratory rhythm unless used at concentrations 10 times larger. In neonates from the monoamine oxidase-A deficient transgenic Tg8 line, SP increased phrenic burst amplitude but had no effect on the respiratory rhythm at the tested concentrations. The role of SP in regulating neonatal respiratory activity is discussed on the basis of rat and mouse results.

In vitro experiments were conducted on neonatal rat brainstem-spinal cord preparations to test the hypothesis of an inhibitory modulation of phrenic activity by serotonin (5-HT) via non-5-HT2A receptors [Lindsay, A.D. and Feldman, J.L., Modulation of respiratory activity of neonatal rat phrenic motoneurones by serotonin, J. Physiol., 461 (1993) 213-233]. The changes induced by 5-HT and related agents on phrenic root discharges and membrane currents in identified phrenic motoneurons were analysed after blockade of spinal 5-HT2A receptors. Spinal application of 5-HT1B (but not 5-HT1A) receptor agonists depressed the phrenic activity and the effect was prevented by pretreatment with 5-HT1B (but not 5-HT1A, 5-HT2A and 5-HT3) receptor antagonists. Results from phrenic motoneuron whole cell recordings do not reject a presynaptic location of the 5-HT receptors responsible for this depression.

The newborn rat brainstem-spinal cord preparation was used to study the effects of tachykinins on the activity of the respiratory rhythm generator in vitro and to characterize the receptors involve. Substance P and tachykinin NK1 and NK3 receptor agonists induced a concentration-dependent increase in the respiratory frequency (10(-9)-10(-7) M), whereas the respiratory frequency was only slightly affected by the tachykinin NK2 receptor agonist. Pre-treatments with tachykinin NK1 receptor antagonists (SR140333, (S)1-¿2-[3-(3.4-dichlorophenyl) -1-(3-isopropoxyphenylacetyl)piperidin-n-3-yl]ethyl¿-4-ph eny l-1-azoniabicyclo [2,2,2]octane chloride; GR82334, pGlu-Ala- Asp-Pro-Asn-Lys-Phe-Tyr-(S-S)Pro-Leu(spiro-gamma-lactam)-Trp-NH2) reduced the substance P-induced increases in the respiratory frequency but the tachykinin NK2 receptor antagonist (SR48968, ((S)-N-methyl-N-[4-4-acetylamino-4-phenylpiperidine)-2-(3,4-dichlorop hen yl) butyl]benzamide); MEN 10376, Asp-Tyr-D-Trp-Val-D-Trp-Lys-NH2) had no effect; the increase in the respiratory frequency induced by the tachykinin NK3 receptor agonist was not affected by a pre-treatment with tachykinin NK1 and NK2 receptor antagonists. These result indicate that tachykinin NK1 and NK3 receptors may be involved in the control of the respiratory frequency.

Ethanol (1-12 mM) added to the superfusion medium of the isolated brainstem-spinal cords of newborn rats did not affect phrenic activity but significantly reduced hypoglossal activity by 54%, 67% and 55% at 3, 6 and 12 mM, respectively. Although the reasons for the suppression of hypoglossal activity remain unknown, this preparation may be a useful model for determining why cranial motoneurons are more vulnerable than phrenic motoneurons to various agents and, more generally, how ethanol impairs neural function.

Previous experiments performed in the in vitro newborn rat brainstem-spinal cord preparation reported that the addition of serotonin (5-HT, 30-50 microM) to the bathing medium induced increases in the respiratory frequency and a large tonic discharge on all the cervical ventral roots. The aim of the present work was to define whether the 5-HT-induced tonic discharge involved respiratory or non-respiratory motoneurones. Intracellular recordings demonstrated that cervical (C2) motoneurones (n = 27) were depolarized by 5-HT but that the 5-HT-induced tonic discharge was mainly due to recruitment of silent motoneurones (n = 18) which fired permanently (15/18; 17 +/- 3 Hz) under 5-HT. The respiratory motoneurones (n = 9) retained a phasic inspiratory discharge (5/9), even if some (4/9) occasionally exhibited a few spikes during expiration. Therefore, it is concluded that the 5-HT-induced tonic discharge is unlikely to have functional significance in respiration.

1. Respiratory activity was recorded on ventral cervical roots during in vitro experiments performed on superfused newborn rat brain stem-cervical cord preparations. 2. Eliminating the pontine structures by performing a transection at the level of the ponto-medullary junction resulted in a sustained increase in respiratory frequency, which suggests the existence of a pontine inhibitory drive impinging on the medullary rhythm generator. 3. Noradrenaline (NA) and drugs affecting NA efficiency were added to the bathing medium and the resulting changes in respiratory frequency were analysed. NA decreased the respiratory frequency, and this effect was potentiated by pargyline (an inhibitor of the NA degradation by monoamine oxidases) and blocked by yohimbine (an alpha 2-antagonist). 4. Yohimbine or piperoxane (which blocks the alpha 2-adrenoceptors) increased the resting respiratory frequency to the level reached after ponto-medullary transection, whereas pargyline or desipramine (which potentiates NA efficiency) decreased the respiratory rate. Since these effects were no longer observed after elimination of the pons, it is suggested that a permanent release of endogenous NA by pontine areas may modulate the activity of the medullary respiratory rhythm generator. 5. When alpha-methyltyrosine (an inhibitor of NA biosynthesis) was applied to the pons, the respiratory frequency was increased, whereas when tyrosine (a precursor of NA) was applied, the respiratory frequency decreased. This decrease was enhanced by pargyline, suppressed by alpha-methyltyrosine and blocked by piperoxane. 6. To conclude, it is suggested that the mechanisms underlying NA biosynthesis (i) continue to function under these in vitro experimental conditions and (ii) are responsible for a permanent release of endogenous NA, which slows down the respiratory frequency. These results are discussed as regards the possibility that the medullary respiratory rhythm generator may be modulated via the noradrenergic area A5 in the newborn rat.

In vitro experiments were performed in the superfused brainstem-spinal cord preparation of newborn rats in order to analyse the central respiratory effects of acetylcholine. The central motor output was assessed from recording electrical activity in nerves supplying respiratory muscles. Acetylcholine added to the bathing medium induced dose-dependent increases in respiratory frequency which were blocked by muscarinic (but not nicotinic) antagonists and enhanced by physostigmine. These effects originated from the medullary ventral surface where chemosensitive structures have been previously located. The respiratory central chemosensitivity of the isolated brainstem was analysed using a CO2 free, pH 7.9 medium instead of the normal medium (bubbled with 5% CO2, pH 7.3). Decreases at the H+ and CO2 stimuli led to decreased inspiratory activity, resulting mainly from a decrease in the amplitude of the motor output. These responses were enhanced by atropine and diminished by physostigmine. These results obtained in vitro on the newborn rat suggest that cholinergic synapses are not directly involved in the genesis of respiratory rhythmicity but confirm previous results obtained in vivo in adult animal revealing that acetylcholine is implicated in the central respiratory chemosensitivity.

Newborn rat respiratory activity was recorded on hypoglossal nerve and ventral cervical roots during in vitro experiments performed on superfused brainstem spinal cord preparations. The addition of serotonin (5-HT) to the bathing medium increased the respiratory frequency and selectively depressed the hypoglossal activity. Any decreases in the amplitude of cervical recordings were always restricted and reversible, whereas the hypoglossal activity was abolished. Furthermore, on cervical roots, 5-HT induced a tonic activity superimposed on the respiratory one, which was never observed with the hypoglossal nerve. When 5-HT was applied on isolated hemispinal cord, a tonic activity could still be elicited. These results indicate that serotonin (i) modulates the activity of neurons involved in the generation of respiratory rhythm, (ii) depresses the activity of hypoglossal motoneurons, and (iii) evokes tonic activity in cervical motoneurons, probably as the result of direct spinal effects.

Abstract: Endogenous cannabinoid receptors are widely distributed throughout the CNS, including the brainstem, and modulate a variety of functions, including breathing. In adult rats, activation of cannabinoid 1 receptors has been shown to depress breathing. Here in neonatal mice, we used in vitro electrophysiology, pharmacology, and immunohistochemistry to analyse the central effects of the endocannabinoid anandamide (AEA) on the activity of the medullary respiratory rhythm generator (RRG). First of all, in vitro electrophysiology on medullary preparations has revealed that bath application of AEA (30 μM, 15 min) significantly depressed respiratory activity. Secondly, applying pre-treatments with alpha-1 (Prazosin, 5 μM, 10 min) and alpha-2 (Yohimbine, 5 μM, 10 min) adrenoceptor antagonists prior to AEA application abolished the AEA-induced depression of the RRG. Finally, immunostaining revealed a dense network of fibres positive for the cannabinoid 1 receptor in the ventrolateral medulla (VLM), a region known to contain both the RRG and the modulatory A1/C1 catecholaminergic group. Moreover, cannabinoid 1 receptor positive fibres were found in close apposition with A1/C1 catecholaminergic cells, identified by the presence of tyrosine hydroxylase. In regard of our electrophysiological, pharmacological and immunostaining results, we conclude that AEA has a central depressive effect on the neonatal RRG, probably via the medullary A1/C1 catecholaminergic neurons which are already known to modulate the respiratory rhythm generator. [ABSTRACT FROM AUTHOR]