Matthew Ireland

Contribution of cholinergic systems to state-dependent modulation of respiratory control.

Respiration is altered during different stages of the sleep-wake cycle. We review the contribution of cholinergic systems to this alteration, with particular reference to the role of muscarinic acetylcholine receptors (MAchRs) during rapid eye movement (REM) sleep. Available evidence demonstrates that MAchRs have potent excitatory effects on medullary respiratory neurones and respiratory motoneurones, and are likely to contribute to changes in central chemosensitive drive to the respiratory control system. These effects are likely to be most prominent during REM sleep, when cholinergic brainstem neurones show peak activity levels. It is possible that MAchR dysfunction is involved in sleep-disordered breathing, such as obstructive sleep apnea.

Mapping the effects of the selective dopamine D2/D3 receptor agonist quinelorane using pharmacological magnetic resonance imaging

Dopamine agonists with a high affinity for D2 and D3 receptors have a biphasic effect on rodent locomotion, inducing hypolocomotion at low doses and hyperlocomotion at higher doses. Controversy surrounds the role of the D3 receptor in mediating the hypolocomotor response to low agonist doses. This study examines patterns of neuronal activation induced by varying doses of the D2/D3 receptor agonist quinelorane using blood oxygen level dependent (BOLD) pharmacological magnetic resonance imaging (phMRI), and compares them with corresponding behavioural responses. Quinelorane (3 microg/kg) induced hypolocomotion in rats naive to the testing environment, and in phMRI experiments increased neuronal activity within the anterior olfactory nuclei, nucleus accumbens and islets of Calleja, regions containing a high density of D3 receptors. A 30 microg/kg dose of quinelorane resulted in biphasic locomotor effects, with initial hypolocomotion followed by sustained hyperlocomotion. phMRI indicated that this higher dose increased cerebral activity within limbic and olfactory regions, as did the lower drug dose, but induced additional activation in the caudate-putamen and globus pallidus, areas dense in D2 receptors but containing few D3 receptors. The more restricted pattern of activation at low agonist doses and close temporal relationship between behavioural and BOLD signal responses to quinelorane suggest that those nuclei most dense in D3 receptors play a key role in mediating the hypolocomotor effects of quinelorane. However, the presence of D3 receptors in activated brain regions may be coincidental, and further studies are required to show definitively which class of receptors mediates agonist-induced hypolocomotion. In contrast, the activation of D2 receptors within the striatum appears necessary for quinelorane-induced hyperlocomotion.

Muscarinic acetylcholine receptors enhance neonatal mouse hypoglossal motoneuron excitability in vitro

In brain stem slices from neonatal (postnatal days 0-4) CD-1 mice, muscarinic ACh receptors (MAChRs) increased rhythmic inspiratory-related and tonic hypoglossal nerve discharge and depolarized single hypoglossal motoneurons (HMs) via an inward current without changing input resistance. These responses were blocked by the MAChR antagonist 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide (4-DAMP; 100 nM). MAChRs shifted voltage-dependent activation of the hyperpolarization-activated cation current to more positive levels. MAChRs increased the HM repetitive firing rate and decreased rheobase, with both effects being blocked by 4-DAMP. Muscarinic agonists reduced the afterhyperpolarization of single action potentials (APs), suggesting that small-conductance Ca(2+)-dependent K(+) current inhibition increased the HM firing rate. Muscarinic agonists also reduced the AP amplitude and slowed its time course, suggesting that MAChRs inhibited voltage-gated Na(+) channels. To compare muscarinic excitation of single HMs to muscarinic excitatory effects on motor output in thicker brain stem slices requiring higher extracellular K(+) for rhythmic activity, we tested the effects of muscarinic agonists on single HM excitability in high-K(+) artificial cerebrospinal fluid (aCSF). In high-K(+) aCSF, muscarinic agonists still depolarized HMs and altered AP size and shape, as in standard aCSF, but did not increase the steady-state firing rate, decrease afterhyperpolarization, or alter threshold potential. These results indicate that the basic cellular response of HMs to muscarinic receptors is excitatory, via a number of distinct mechanisms, and that this excitatory response will be largely preserved in rhythmically active brain stem slices.

Distinct receptors underlie glutamatergic signalling in inspiratory rhythm-generating networks and motor output pathways in neonatal rat

Despite the enormous diversity of glutamate (Glu) receptors and advances in understanding recombinant receptors, native Glu receptors underlying functionally identified inputs in active systems are poorly defined in comparison. In the present study we use UBP‐302, which antagonizes GluR5 subunit‐containing kainate (KA) receptors at ≤ 10 μm, but other KA and AMPA receptors at ≥ 100 μm, and rhythmically active in vitro preparations of neonatal rat to explore the contribution of non‐NMDA receptor signalling in rhythm‐generating and motor output compartments of the inspiratory network. At 10 μm, UBP‐302 had no effect on inspiratory burst frequency or amplitude. At 100 μm, burst amplitude recorded from XII, C1 and C4 nerve roots was significantly reduced, but frequency was unaffected. The lack of a frequency effect was confirmed when local application of UBP‐302 (100 μm) into the pre‐Bötzinger complex (preBötC) did not affect frequency but substance P evoked a 2‐fold increase. A UBP‐302‐sensitive (10 μm), ATPA‐evoked frequency increase, however, established that preBötC networks are sensitive to GluR5 activation. Whole‐cell recordings demonstrated that XII motoneurons also express functional GluR5‐containing KA receptors that do not contribute to inspiratory drive, and confirmed the dose dependence of UBP‐302 actions on KA and AMPA receptors. Our data provide the first evidence that the non‐NMDA (most probably AMPA) receptors mediating glutamatergic transmission within preBötC inspiratory rhythm‐generating networks are pharmacologically distinct from those transmitting drive to inspiratory motoneurons. This differential expression may ultimately be exploited pharmacologically to separately counteract depression of central respiratory rhythmogenesis or manipulate the drive to motoneurons controlling airway and pump musculature.

Contribution of cholinergic systems to state-dependent modulation of respiratory networks

The sleep/wake cycle is controlled by reciprocal inhibition between cholinergic and aminergic cell groups in the pons and brainstem [1]. In rapid eye movement (REM) sleep, cholinergic neurone discharge is at its highest level while aminergic discharge is at the lowest level across all behavioural states [1]. These changes are accompanied by alteration of respiratory pattern and in the output of some respiratory motoneurone pools [1]. While effects of aminergic receptors on respiratory pattern and motor output have been extensively investigated, less is known of the effects of cholinergic receptors. Disease syndromes such as obstructive sleep apnoea in adults or infants, or sudden infant death syndrome may involve abnormal cholinergic receptor-mediated responses. Here we report network and cellular effects of muscarinic acetylcholine receptors (mAChRs) on respiratory pattern and motor output in hypoglossal motoneurons (HMs) using in vitro slice preparations from mouse. Rhythmically active brainstem slices were made from mice (P0-4 days) and the mAChR agonist, muscarine was bath applied (10 µM, n = 6). As illustrated in Fig. ​Fig.1A,1A, integrated hypoglossal nerve (IntXIIN) burst amplitude increased by 80 ± 24% (mean ± SE) while burst frequency decreased (-34 ± 7%). Local injection of muscarine (100 µM, n = 14) over the hypoglossal nucleus (nXII) produced an increase in IntXIIN burst amplitude (96 ± 23%) but no change in burst frequency (5 ± 4%). Injections of muscarine (n = 3) in the pre-Botzinger complex (PBC) decreased IntXIIN burst frequency (-20 ± 5%) and increased burst amplitude (60 ± 12%). These effects were blocked by bath application of atropine (n = 3, 1 µM). In whole cell recordings from HMs (n = 14) local nXII or bath application of muscarine evoked an inward current (-29 ± 5 pA) in voltage clamp at -60 mV (see Fig. ​Fig.1A1A for example) and depolarization in current clamp. This current was associated with increased neuronal input resistance (Rn) which was greater at voltages positive to -60 mV (Fig. ​(Fig.1B),1B), suggesting that muscarine inhibited a voltage-dependent outward current. Muscarinic inhibition of K+ currents elicited by slow (2 s) depolarizing voltage ramps in the presence of TTX and Cd2+ was much greater at voltages from -70 to -20 mV (Fig. ​(Fig.1C),1C), a voltage range typically occupied by the voltage- and ligand-dependent K+current IM but not by other voltage-dependent K+ currents. Figure 1 These results suggest that mAChRs have potent effects both on respiratory rhythm generation and HM motor output by distinct actions on both PBC neurones and HMs. We hypothesize that excitatory effects of mAChRs on HMs is partly due to muscarinic inhibition of the potassium current IM, causing depolarization and increased Rn and thus enhancing HM firing.