Excitability of Inspiratory and Non-Inspiratory Hypoglossal Motoneurons is Increased by Muscarinic Modulation in Neonatal Mice
Authors List
Ann L Revill1, Sydney K Dudley1 & Sarena Fernandez2
1Department of Physiology, College of Graduate Studies; 2Arizona College of Osteopathic Medicine, Midwestern University, Glendale, Arizona
Hypoglossal motoneurons (XII MNs) that innervate the tongue decrease activity normally during sleep, which leads to a reduction in airway tone. The specific mechanisms that contribute to decreased airway tone during sleep remain to be completely resolved, although research from adult rodents suggests activation of muscarinic acetylcholine receptors (mAChRs) is an important contributor by inhibiting XII MNs through either pre- or post-synaptic mechanisms. Conversely, data from neonatal rodents, have shown that muscarinic modulation leads to an overall excitatory effect in XII MNs. As such, the overarching objective of this research project is to uncover the mechanisms through which muscarinic modulation can influence the excitability of XII MNs across postnatal maturation. We hypothesized that mAChR activation at XII MNs would increase excitability of non-inspiratory and inspiratory XII MNs, as well as potentiate inspiratory bursting in inspiratory XII MNs. We first tested local application of muscarine (100 μM) into the hypoglossal nucleus using the in-vitro rhythmic slice preparation (CD1 mice) from postnatal day (P) 0 to 14. We then used rhythmic medullary slices from neonatal mice (postnatal day 0-5) for whole cell electrophysiology experiments to test the effects of activating mAChRs at XII MNs in both non-inspiratory and inspiratory XII MNs. Data are reported as mean ± SD.
Inspiratory burst amplitude was potentiated by muscarine from postnatal (P) 0-11 although preliminary data suggest muscarinic modulation attenuated inspiratory burst amplitude at P14. Fitting the data with a polynomial, the magnitude of potentiation appeared to change after P4, P9 and P14, indicating that age influences muscarinic potentiation (p=0.0071, n=16). For whole cell experiments with XII MNs clamped at -60 mV, muscarinic modulation depolarized non-inspiratory and inspiratory XII MNs similarly (3.6±2.6 vs 4.6±2.1 mV, n=12, p>0.05), and had no effect on input resistance (CTL vs Musc; non-inspiratory: 120±23 vs 128 ±33 MW; inspiratory: 140±28 vs 137±32 MW; n=12; p>0.05). Muscarinic modulation significantly increased the firing frequency after muscarine application for inspiratory cells with the current step injections 100 pA (CTL vs Musc, 14.7±6.8 vs 21.1±6.4 Hz, n=9 and 10, p<0.05) and 175 pA (CTL vs Musc; 19.5±6.4 vs 25.0±9.4 Hz, n=11 and 8; p<0.05). Finally, muscarinic modulation significantly increased inspiratory burst amplitude (CTL vs Musc; 15.2±3.1 vs 18.3±4.5 mV; n=3; p<0.05), and a trend to increase inspiratory burst area (CTL vs Musc; 4260±779 vs 5354±306 mV.ms, n=3; p=0.08).
These data indicate that muscarinic modulation has similar effects on non-inspiratory and inspiratory XII MNs early in postnatal maturation, although the magnitude of this effect varies with postnatal age.
Funding: NIH R15 R15HL148870 to ALR.
Ann L Revill1, Sydney K Dudley1 & Sarena Fernandez2
1Department of Physiology, College of Graduate Studies; 2Arizona College of Osteopathic Medicine, Midwestern University, Glendale, Arizona
Hypoglossal motoneurons (XII MNs) that innervate the tongue decrease activity normally during sleep, which leads to a reduction in airway tone. The specific mechanisms that contribute to decreased airway tone during sleep remain to be completely resolved, although research from adult rodents suggests activation of muscarinic acetylcholine receptors (mAChRs) is an important contributor by inhibiting XII MNs through either pre- or post-synaptic mechanisms. Conversely, data from neonatal rodents, have shown that muscarinic modulation leads to an overall excitatory effect in XII MNs. As such, the overarching objective of this research project is to uncover the mechanisms through which muscarinic modulation can influence the excitability of XII MNs across postnatal maturation. We hypothesized that mAChR activation at XII MNs would increase excitability of non-inspiratory and inspiratory XII MNs, as well as potentiate inspiratory bursting in inspiratory XII MNs. We first tested local application of muscarine (100 μM) into the hypoglossal nucleus using the in-vitro rhythmic slice preparation (CD1 mice) from postnatal day (P) 0 to 14. We then used rhythmic medullary slices from neonatal mice (postnatal day 0-5) for whole cell electrophysiology experiments to test the effects of activating mAChRs at XII MNs in both non-inspiratory and inspiratory XII MNs. Data are reported as mean ± SD.
Inspiratory burst amplitude was potentiated by muscarine from postnatal (P) 0-11 although preliminary data suggest muscarinic modulation attenuated inspiratory burst amplitude at P14. Fitting the data with a polynomial, the magnitude of potentiation appeared to change after P4, P9 and P14, indicating that age influences muscarinic potentiation (p=0.0071, n=16). For whole cell experiments with XII MNs clamped at -60 mV, muscarinic modulation depolarized non-inspiratory and inspiratory XII MNs similarly (3.6±2.6 vs 4.6±2.1 mV, n=12, p>0.05), and had no effect on input resistance (CTL vs Musc; non-inspiratory: 120±23 vs 128 ±33 MW; inspiratory: 140±28 vs 137±32 MW; n=12; p>0.05). Muscarinic modulation significantly increased the firing frequency after muscarine application for inspiratory cells with the current step injections 100 pA (CTL vs Musc, 14.7±6.8 vs 21.1±6.4 Hz, n=9 and 10, p<0.05) and 175 pA (CTL vs Musc; 19.5±6.4 vs 25.0±9.4 Hz, n=11 and 8; p<0.05). Finally, muscarinic modulation significantly increased inspiratory burst amplitude (CTL vs Musc; 15.2±3.1 vs 18.3±4.5 mV; n=3; p<0.05), and a trend to increase inspiratory burst area (CTL vs Musc; 4260±779 vs 5354±306 mV.ms, n=3; p=0.08).
These data indicate that muscarinic modulation has similar effects on non-inspiratory and inspiratory XII MNs early in postnatal maturation, although the magnitude of this effect varies with postnatal age.
Funding: NIH R15 R15HL148870 to ALR.
