Mechanisms underlying the endogenous dopaminergic inhibition of spinal locomotor circuit function in Xenopus tadpoles

This work was supported by the Biotechnology and Biological Science Research Council (BBSRC) [grant number BB/J01446X/1].Dopamine plays important roles in the development and modulation of motor control circuits. Here we show that dopamine exerts potent effects on the central pattern generator circuit controlling locomotory swimming in post-embryonic Xenopus tadpoles. Dopamine (0.5–100 μM) reduced fictive swim bout occurrence and caused both spontaneous and evoked episodes to become shorter, slower and weaker. The D2-like receptor agonist quinpirole mimicked this repertoire of inhibitory effects on swimming, whilst the D4 receptor antagonist, L745,870, had the opposite effects. The dopamine reuptake inhibitor bupropion potently inhibited fictive swimming, demonstrating that dopamine constitutes an endogenous modulatory system. Both dopamine and quinpirole also inhibited swimming in spinalised preparations, suggesting spinally located dopamine receptors. Dopamine and quinpirole hyperpolarised identified rhythmically active spinal neurons, increased rheobase and reduced spike probability both during swimming and in response to current injection. The hyperpolarisation was TTX-resistant and was accompanied by decreased input resistance, suggesting that dopamine opens a K+ channel. The K+ channel blocker barium chloride (but not TEA, glybenclamide or tertiapin-Q) significantly occluded the hyperpolarisation. Overall, we show that endogenously released dopamine acts upon spinally located D2-like receptors, leading to a rapid inhibitory modulation of swimming via the opening of a K+ channel.Publisher PDFPeer reviewe

The roles of dopamine and the sodium pump in the spinal control of locomotion

Rhythmically active, locomotor networks of the spinal cord are subject to both neuromodulation and activity-dependent homeostatic regulation. I first show that the neuromodulator dopamine exerts potent inhibitory effects on the central pattern generator (CPG) circuit controlling locomotory swimming in post-embryonic Xenopus tadpoles. Dopamine, acting endogenously on spinal D2-like receptors, reduces spontaneous fictive swimming occurrence and shortens, slows and weakens swimming. The mechanism involves a TTX-resistant hyperpolarisation of rhythmically active CPG neurons, mediated by the direct opening of a K+ channel with GIRK-like pharmacology. This increases rheobase and reduces spike probability. I next explore how sodium pumps contribute to the activity-dependent regulation of the Xenopus swim circuit, and possible interactions of the pumps with modulators, temperature and ionic conductances. I characterise the pump-mediated ultra-slow afterhyperpolarisation (usAHP), and show that monensin, a sodium ionophore, enhances pump activity, converting the usAHP into a tonic hyperpolarisation; this decreases swim episode duration and cycle frequency. I also characterise a ZD7288-sensitive Ih current, which is active in excitatory dIN interneurons and contributes to spiking. Blocking Ih with ZD7288 decreases swim episode duration and destabilises swim bursts. Both Ih and the usAHP increase with temperature, which depolarises CPG neurons, decreases input resistance, and increases spike probability; this increases cycle frequency, but the enhanced usAHP shortens swimming. I also show that the usAHP is diminished by nitric oxide, but enhanced by dopaminergic signalling. Finally, I explore sodium pumps in the neonatal mouse. The sodium pump blocker ouabain increases the duration and frequency of drug- and sensory-induced locomotion, whilst monensin has opposite effects. Decreasing inter-episode interval also shortens and slows activity, a relationship abolished by ouabain, implicating sodium pumps in a feedforward motor memory mechanism. Finally, I show that the effects of ouabain on locomotion are dependent on dopamine, which enhances a TTX- and ouabain-sensitive usAHP in spinal neurons

Control of <i>Xenopus</i> tadpole locomotion via selective expression of Ih in excitatory interneurons

The authors are grateful for the support of the Biotechnology and Biological Science Research Council (BBSRC) [grant number BB/J01446X/1 and BB/M024946/1] and the Wellcome Trust-University of Edinburgh Institutional Strategic Support Fund (ISSF).Locomotion relies on the coordinated activity of rhythmic neurons in the hindbrain and spinal cord, and depends critically on the intrinsic properties of excitatory interneurons. Therefore, understanding how ion channels sculpt the properties of these interneurons, and the consequences for circuit function and behavior, is an important task. The hyperpolarization-activated cation current, Ih, is known to play important roles in shaping neuronal properties and for rhythm generation in many neuronal networks. We show in stage 42 Xenopus laevis frog tadpoles that Ih is strongly expressed only in excitatory descending interneurons (dINs), an important ipsilaterally projecting population that drives swimming activity. The voltage-dependent HCN channel blocker ZD7288 completely abolished a prominent depolarising sag potential in response to hyperpolarization, the hallmark of Ih, and hyperpolarized dINs. ZD7288 also affected dIN post-inhibitory rebound firing, upon which locomotor rhythm generation relies, and disrupted locomotor output. Block of Ih also unmasked an activity-dependent ultraslow afterhyperpolarization (usAHP) in dINs following swimming, mediated by a dynamic Na/K pump current. This usAHP, unmasked in dINs by ZD7288, resulted in suprathreshold stimuli failing to evoke swimming at short inter-swim intervals, indicating an important role for Ih in maintaining swim generation capacity and in setting the post-swim refractory period of the network. Collectively, our data suggest that the selective expression of Ih in dINs determines specific dIN properties that are important for rhythm generation and counteracts an activity- dependent usAHP to ensure that dINs can maintain coordinated swimming over a wide range of inter-swim intervals.Publisher PDFPeer reviewe

Sodium pump regulation of locomotor control circuits

The authors are grateful for the financial support of the BBSRC (grant numbers: BB/M024946/1 and BB/JO1446X/1), the Carnegie Trust and the University of St Andrews.Sodium pumps are ubiquitously expressed membrane proteins that extrude three Na+ ions in exchange for two K+ ions using ATP as an energy source. Recent studies have illuminated additional, dynamic roles for sodium pumps in regulating the excitability of neuronal networks in an activity-dependent fashion. Here we review their role in a novel form of short-term memory within rhythmic locomotor networks. The data we review derives mainly from recent studies on Xenopus tadpoles and neonatal mice. The role and underlying mechanisms of pump action broadly match previously published data from an invertebrate, the Drosophila larva. We therefore propose a highly conserved mechanism by which sodium pump activity increases following a bout of locomotion. This results in an ultraslow afterhyperpolarisation (usAHP) of the membrane potential that lasts around 1 minute, but which only occurs in around half the network neurons. This usAHP in turn alters network excitability so that network output is reduced in a locomotor interval-dependent manner. The pumps therefore confer on spinal locomotor networks a temporary memory trace of recent network performance.PostprintPeer reviewe

Bimodal modulation of short-term motor memory via dynamic sodium pumps in a vertebrate spinal cord

Authors are grateful for the financial support of this research by the University of St Andrews, the E&RS Neuroscience research fund and BBSRC grant BB/T015705/1.Dynamic neuronal Na+/K+ pumps normally only respond to intense action potential firing owing to their low affinity for intracellular Na+. Recruitment of these Na+ pumps produces a post-activity ultraslow afterhyperpolarization (usAHP) up to ∼10 mV in amplitude and ∼60 s in duration, which influences neuronal properties and future network output. In spinal motor networks, the usAHP underlies short-term motor memory (STMM), reducing the intensity and duration of locomotor network output in a manner dependent on the interval between locomotor bouts. In contrast to tonically active Na+ pumps that help set and maintain the resting membrane potential, dynamic Na+ pumps are selectively antagonized by low concentrations of ouabain, which, we show, blocks both the usAHP and STMM. We examined whether dynamic Na+ pumps and STMM can be influenced by neuromodulators, focusing on 5-HT and nitric oxide. Bath-applied 5-HT alone had no significant effect on the usAHP or STMM. However, this is due to the simultaneous activation of two distinct 5-HT receptor subtypes (5-HT7 and 5-HT2a) that have opposing facilitatory and suppressive influences, respectively, on these two features of the locomotor system. Nitric oxide modulation exerts a potent inhibitory effect that can completely block the usAHP and erase STMM. Using selective blockers of 5-HT7 and 5-HT2a receptors and a nitric oxide scavenger, PTIO, we further provide evidence that the two modulators constitute an endogenous control system that determines how the spinal network self-regulates the intensity of locomotor output in light of recent past experience.Publisher PDFPeer reviewe

Sodium pumps mediate activity-dependent changes in mammalian motor networks

Funding: Laurence Picton [grant number BB/JO1446X/1] and Matthew Broadhead [grant number BB/M021793/1] were supported by the Biotechnology and Biological Science Research Council (BBSRC). Filipe Nascimento was supported by The Alfred Dunhill Links Foundation.Ubiquitously expressed sodium pumps are best known for maintaining the ionic gradients and resting membrane potential required for generating action potentials. However, activity- and state-dependent changes in pump activity can also influence neuronal firing and regulate rhythmic network output. Here we demonstrate that changes in sodium pump activity regulate locomotor networks in the spinal cord of neonatal mice. The sodium pump inhibitor, ouabain, increased the frequency and decreased the amplitude of drug-induced locomotor bursting, effects that were dependent on the presence of the neuromodulator dopamine. Conversely, activating the pump with the sodium ionophore monensin decreased burst frequency. When more "natural" locomotor output was evoked using dorsal-root stimulation, ouabain increased burst frequency and extended locomotor episode duration, whereas monensin slowed and shortened episodes. Decreasing the time between dorsal-root stimulation, and therefore interepisode interval, also shortened and slowed activity, suggesting that pump activity encodes information about past network output and contributes to feedforward control of subsequent locomotor bouts. Using whole-cell patch-clamp recordings from spinal motoneurons and interneurons, we describe a long-duration (∼60 s), activity-dependent, TTX- and ouabain-sensitive, hyperpolarization (∼5 mV), which is mediated by spike-dependent increases in pump activity. The duration of this dynamic pump potential is enhanced by dopamine. Our results therefore reveal sodium pumps as dynamic regulators of mammalian spinal motor networks that can also be affected by neuromodulatory systems. Given the involvement of sodium pumps in movement disorders, such as amyotrophic lateral sclerosis and rapid-onset dystonia parkinsonism, knowledge of their contribution to motor network regulation also has considerable clinical importance.Publisher PDFPeer reviewe

Nouveaux systèmes micellaires intelligents à partir d'huile de lin (synthèse, comportements physico-chimiques et encapsulation)

Les micelles apparaissent comme prometteuses dans le domaine de la vectorisation de médicaments. Afin d améliorer leur biocompatibilité nous nous intéressons ici à des synthèses originales de copolymères amphiphiles comportant un bloc hydrophobe lipidique biosourcé. Un polymère intelligent constitue le bloc hydrophile. L huile de lin subit une modification chimique afin d introduire un site amorceur de polymérisation. Le bloc hydrophile est alors ajouté par polymérisation contrôlée. Deux copolymères sont obtenus, le lipide-b-poly(acide acrylique), pH-sensible et le lipide-b-poly(2-isopropyl-oxazoline), thermo-sensible. Une caractérisation physico-chimique complète révèle des concentrations micellaires critiques basses et des micelles de 10 nm. Un système mixte est préparé par mélange des deux copolymères. L étude de ce système prouve une sensibilité aux deux stimuli. Afin d améliorer la stabilité des micelles, nous proposons la photo-réticulation des insaturations de la chaîne lipidique.Small micellar systems seem to be really promising candidates for drug delivery applications. In order to improve the bio-assimilation of our system, the original synthesis of amphiphilic copolymers from linseed oil is carried out. First, linseed oil is chemically modified in order to introduce a polymerization initiating site. Then, the lipoinitiator is engaged in the controlled polymerization of the hydrophilic block. Two amphiphilic copolymers are obtained through this strategy: a pH-sensitive lipid-b-poly(acrylic acid), and a thermo-sensitive lipid-b-poly(2-isopropyl-oxazoline). Both present a low critical micellar concentration and form small micelles (~10 nm). By mixing both copolymers, mixed micelles responding to both stimuli were obtained. In order to improve the system s stability, the photo-cross-linking of the lipidic double bonds in the micelle s core is finally realized.ROUEN-INSA Madrillet (765752301) / SudocSudocFranceF

Mechanisms underlying the activity-dependent regulation of locomotor network performance by the Na⁺ pump

This research was funded by BBSRC (project grants to KTS (BB/F015488/1) and W-CL; EastBio studentship to LP) and Wellcome Trust (ISSF to H-YZ).Activity-dependent modification of neural network output usually results from changes in neurotransmitter release and/or membrane conductance. In Xenopus frog tadpoles, spinal locomotor network output is adapted by an ultraslow afterhyperpolarization (usAHP) mediated by an increase in Na+ pump current. Here we systematically explore how the interval between two swimming episodes affects the second episode, which is shorter and slower than the first episode. We find the firing reliability of spinal rhythmic neurons to be lower in the second episode, except for excitatory descending interneurons (dINs). The sodium/proton antiporter, monensin, which potentiates Na+ pump function, induced similar effects to short inter-swim intervals. A usAHP induced by supra-threshold pulses reduced neuronal firing reliability during swimming. It also increased the threshold current for spiking and introduced a delay to the first spike in a train, without reducing subsequent firing frequency. This delay was abolished by ouabain or zero K+ saline, which eliminate the usAHP. We present evidence for an A-type K+ current in spinal CPG neurons which is inactivated by depolarization and de-inactivated by hyperpolarization, and accounts for the prolonged delay. We conclude that the usAHP attenuates neuronal responses to excitatory network inputs by both membrane hyperpolarization and enhanced de-inactivation of an A-current.Publisher PDFPeer reviewe

Principles Governing Locomotion in Vertebrates: Lessons From Zebrafish

Locomotor behaviors are critical for survival and enable animals to navigate their environment, find food and evade predators. The circuits in the brain and spinal cord that initiate and maintain such different modes of locomotion in vertebrates have been studied in numerous species for over a century. In recent decades, the zebrafish has emerged as one of the main model systems for the study of locomotion, owing to its experimental amenability, and work in zebrafish has revealed numerous new insights into locomotor circuit function. Here, we review the literature that has led to our current understanding of the neural circuits controlling swimming and escape in zebrafish. We highlight recent studies that have enriched our comprehension of key topics, such as the interactions between premotor excitatory interneurons (INs) and motoneurons (MNs), supraspinal and spinal circuits that coordinate escape maneuvers, and developmental changes in overall circuit composition. We also discuss roles for neuromodulators and sensory inputs in modifying the relative strengths of constituent circuit components to provide flexibility in zebrafish behavior, allowing the animal to accommodate changes in the environment. We aim to provide a coherent framework for understanding the circuitry in the brain and spinal cord of zebrafish that allows the animal to flexibly transition between different speeds, and modes, of locomotion