Hypoglossal motoneurons (XII MNs) control muscles of the mammalian tongue and are rhythmically active during breathing. Acetylcholine (ACh) modulates XII MN activity by promoting the release of glutamate from neurons that express nicotinic ACh receptors (nAChRs). Chronic nicotine exposure alters nAChRs on neurons throughout the brain, including brain stem respiratory neurons. Here we test the hypothesis that developmental nicotine exposure (DNE) reduces excitatory synaptic input to XII MNs. Voltage-clamp experiments in rhythmically active medullary slices showed that the frequency of excitatory postsynaptic currents (EPSCs) onto XII MNs from DNE animals is reduced by 61% (DNE = 1.7 ± 0.4 events/s; control = 4.4 ± 0.6 events/s; P < 0.002). We also examine the intrinsic excitability of XII MNs to test whether cells from DNE animals have altered membrane properties. Current-clamp experiments showed XII MNs from DNE animals had higher intrinsic excitability, as evaluated by measuring their response to injected current. DNE cells had high-input resistances (DNE = 131.9 ± 13.7 MΩ, control = 78.6 ± 9.7 MΩ, P < 0.008), began firing at lower current levels (DNE = 144 ± 22 pA, control = 351 ± 45 pA, P < 0.003), and exhibited higher frequency-current gain values (DNE = 0.087 ± 0.012 Hz/pA, control = 0.050 ± 0.004 Hz/pA, P < 0.02). Taken together, our data show previously unreported effects of DNE on XII MN function and may also help to explain the association between DNE and the incidence of central and obstructive apneas.

Pilarski JQ, Wakefield HE, Fuglevand AJ, Levine RB, Fregosi RF. Increased nicotinic receptor desensitization in hypoglossal motor neurons following chronic developmental nicotine exposure. J Neurophysiol 107: 257-264, 2012. First published October 19, 2011 doi:10.1152/jn.00623.2011.-Neuronal nicotinic acetylcholine receptors (nAChRs) are expressed on hypoglossal motor neurons (XII MNs) that innervate muscles of the tongue. Activation of XII MN nAChRs evokes depolarizing currents, which are important for regulating the size and stiffness of the upper airway. Although data show that chronic developmental nicotine exposure (DNE) blunts cholinergic neurotransmission in the XII motor nucleus, it is unclear how nAChRs are involved. Therefore, XII MN nAChR desensitization and recovery were examined in tissues from DNE or control pups using a medullary slice preparation and tight-seal whole cell patch-clamp recordings. nAChR-mediated inward currents were evoked by brief pressure pulses of nicotine or the ␣4␤2 nAChR agonist RJR-2403. We found that, regardless of treatment, activatable nAChRs underwent desensitization, but, following DNE, nAChRs exhibited increased desensitization and delayed recovery. Similar results were produced using RJR-2403, showing that DNE influences primarily the ␣4␤2 nAChR subtype. These results show that while some nAChRs preserve their responsiveness to acute nicotine following DNE, they more readily desensitize and recover more slowly from the desensitized state. These data provide new evidence that chronic DNE modulates XII MN nAChR function, and suggests an explanation for the association between DNE and the incidence of central and obstructive apneas.

The maximum firing rates of motoneurons (MNs), activated in response to synaptic drive, appear to be much lower than that elicited by current injection. It could be that the decrease in input resistance associated with increased synaptic activity (but not current injection) might blunt overall changes in membrane depolarization and thereby limit spike-frequency output. To test this idea, we recorded, in the same cells, maximal firing responses to current injection and to synaptic activation. We prepared 300 μm medullary slices in neonatal rats that contained hypoglossal MNs and used whole-cell patch-clamp electrophysiology to record their maximum firing rates in response to triangular-ramp current injections and to glutamate receptor-mediated excitation. Brief pressure pulses of high-concentration glutamate led to significant depolarization, high firing rates, and temporary cessation of spiking due to spike inactivation. In the same cells, we applied current clamp protocols that approximated the time course of membrane potential change associated with glutamate application and with peak current levels large enough to cause spike inactivation. Means (SD) of maximum firing rates obtained in response to glutamate application were nearly identical to those obtained in response to ramp current injection [glutamate 47.1 ± 12.0 impulses (imp)/s, current injection 47.5 ± 11.2 imp/s], even though input resistance was 40% less during glutamate application compared with current injection. Therefore, these data suggest that the reduction in input resistance associated with receptor-mediated excitation does not, by itself, limit the maximal firing rate responses in MNs.

Although nicotinic acetylcholine receptors (nAChRs) are widespread in the CNS their role in the control of breathing is still unclear. This study aims to test the hypothesis that alterations in nAChRs, evoked by chronic nAChR excitation, impairs respiratory‐related neurotransmission onto hypoglossal motoneurons (HMN) that control the tongue. To test our hypothesis we used rhythmic medullary slice preparations and simultaneously recorded from both single respiratory modulated HMN and whole hypoglossal nerve rootlets on day 1–4 in rat pups chronically exposed to nicotine or saline in utero and during the first week of life. Results show that synaptic strength, predominately glutamatergic, was decreased but without changes in the frequency of respiratory rhythm determined by the respiratory central pattern generator (CPG). The average number of spontaneous excitatory postsynaptic potentials (EPSPs), measured during the “expiratory” period, was decreased 42% (P < 0.05), but their magnitude was unchanged. Furthermore, EPSPs could be abolished by bath application of AMPAergic antagonist 6‐cyano‐7‐nitroquinoxaline‐2, 3‐dione (CNQX). These results show that the chronic in utero exposure to nicotine decreases excitatory neurotransmission at HMN synapses and suggests that this change results from frequency of nAChR‐mediated presynpatic release of glutamate. Funded by NIH #1R03HD061613‐01.

Hypoglossal motor neurons (HMNs) participate in a host of oromotor behaviors including swallowing and breathing. Although these neurons receive input from diverse neurotransmitter systems, of critical importance is the central cholinergic system, which participates in chemosensitivity and state changes. Prenatal nicotine exposure (PNE) reduces efficacy of nicotinic acetylcholine receptors and is associated with a variety of neonatal respiratory abnormalities. Therefore, the purpose of this study was to determine the effects of PNE on intrinsic properties of HMNs which mediate respiratory output to tongue muscles. We obtained rhythmic medullary slice preparations from P1–P4 rats exposed to saline (n = 6) or nicotine (n = 5) in utero, and we recorded from HMNs using whole cell patch clamp electrophysiology. Based on current clamp protocols applied in between respiratory bursts, we found that HMNs of PNE animals had altered firing profiles compared to HMNs of control animals. Specifically, PNE cells required 31% less current to elicit firing and had 49% higher initial frequency‐current gain values compared to control. This change in responsiveness cannot be explained by differences in input resistance. These data suggest that PNE augments HMN intrinsic excitability, which could be a result of neuroplasticity caused by chronic nicotine exposure. This study was funded by American Heart Association #0855713G.