Electrophysiological investigations of the anterior and posterior lateral line nerve of the Goldfish, Carassius auratus, to running water and oscillatory stimuli

The neural responses of anterior lateral line nerve (ALLN) fibers of goldfish to sinusoidal water motions in still and running water were investigated. In agreement with previous data (Engelmann et al., 2002) two types of fibers were distinguished: type I fibers, which most likely innervate superficial neuromasts, were stimulated by running water (10 cm/s) and, type II fibers, which most likely innervate canal neuromasts, were not stimulated by running water. The responses of type I fibers to sinusoidal water motions were masked in running water whereas responses of type II fibers were not masked. The degree of response masking increased with increasing flow velocity. Moreover, the ratio between responses that were masked in running water (type I) and those that were not masked (type II) increased with increasing flow velocity. Fibers exposed to unidirectional water flow showed a continuum of flow sensitivity and not two separate populations of flow sensitive and flow insensitive fibers as might be expected from previous results and theoretical considerations. Most nerve fibers responded with an increase in discharge rate, irrespective of flow direction (head-to-tail and tail-to-head flow). Thus fibers showed no directional sensitivity. Frequency spectra of water motions quantified with particle image velocimetry (PIV) and spectra of the firing rate of lateral line fibers showed an increase in amplitude below 10 Hz under flow conditions. This suggests that the neuromasts responded to the flow fluctuations that developed under flow conditions, but not to the d.c. flow. Thus it is unlikely that the spike trains of individual fibers code for the direction and velocity of a constant flow. Spike trains of ALLN fibers stimulated by a Kármán vortex street (KVS) showed that low frequency stimuli can still be encoded under running water. In terms of spike rate there was no difference between the KVS and the running water condition. However, if exposed to vortex stimuli, spike train frequency spectra showed reproducible peaks at the vortex shedding frequency. Any change in the vortex shedding frequency evoked by a change in either cylinder diameter or water flow velocity shifted the reproducible peaks of the neuronal data into the expected direction. Thus the data show that information about the frequency composition of flow fluctuation is preserved in the spike trains under flow conditions despite the fact that the fibers do not code for the direction and velocity of the flow

Vocal behavior and vocal central pattern generator organization diverge among toadfishes

Among fishes, acoustic communication is best studied in toadfishes, a single order and family that includes species commonly known as toadfish and midshipman. However, there is a lack of comparative anatomical and physiological studies, making it difficult to identify both shared and derived mechanisms of vocalization among toadfishes. Here, vocal nerve labeling and intracellular in vivo recording and staining delineated the hindbrain vocal network of the Gulf toadfish Opsanus beta. Dextran-biotin labeling of the vocal nerve or intracellular neurobiotin fills of motoneurons delineated a midline vocal motor nucleus (VMN). Motoneurons showed bilaterally extensive dendritic arbors both within and lateral to the paired motor nuclei. The motoneuron activity matched that of the spike-like vocal nerve motor volley that determines the natural call duration and frequency. Ipsilateral vocal nerve labeling with biocytin or neurobiotin yielded dense bilateral transneuronal filling of motoneurons and coextensive columns of premotor neurons. These premotor neurons generated pacemaker-like action potentials matched 1:1 with vocal nerve and motoneuron firing. Transneuronal transport further revealed connectivity within and between the pacemaker-motor circuit and a rostral prepacemaker nucleus. Unlike the pacemaker-motor circuit, prepacemaker firing did not match the frequency of vocal nerve activity but instead was predictive of the duration of the vocal nerve volley that codes for call duration. Transneuronally labeled terminal-like boutons also occurred in auditory-recipient hindbrain nuclei, including neurons innervating the inner ear and lateral line organs. Together with studies of midshipman, we propose that separate premotor populations coding vocal frequency and duration with direct premotor coupling to auditory-lateral line nuclei are plesiomorphic characters for toadfishes. Unlike in midshipman, transneuronal labeling in toadfishes reveals an expansive column of pacemaker neurons that is weakly coupled to prepacemaker neurons, a character that likely depends on the extent of gap junction coupling. We propose that these and other anatomical characters contribute to neurophysiological properties that, in turn, sculpt the species-typical patterning of frequency and amplitude-modulated vocalizations

Serotonin systems in three socially communicating teleost species, the grunting toadfish (Allenbatrachus grunniens), a South American marine catfish (Ariopsis seemanni), and the upside-down catfish (Synodontis nigriventris)

We investigated immunohistochemically the distribution of serotonergic cell populations in three teleost species (one toadfish, Allenbatrachus grunniens, and two catfishes, Synodontis nigriventris and Ariopsis seemanni). All three species exhibited large populations of 5-HT positive neurons in the paraventricular organ (PVO) and the dorsal (Hd) and caudal (Hc) periventricular hypothalamic zones, plus a smaller one in the periventricular pretectum, a few cells in the pineal stalk, and – only in catfishes – in the preoptic region. Furthermore, the rhombencephalic superior and inferior raphe always contained ample serotonergic cells. In each species, a neuronal mass extended into the hypothalamic lateral recess. Only in the toadfish, did this intraventricular structure contain serotonergic cells and arise from Hd, whereas in the catfishes it emerged from medially and represents the dorsal tuberal nucleus seen in other catfishes as well. Serotonergic cells in PVO, Hd and Hc were liquor-contacting. Those of the PVO extended into the midline area of the periventricular posterior tubercular nucleus in both catfishes. Dopaminergic, liquor-contacting neurons were additionally investigated using an antibody against tyrosine hydroxylase (TH) in S. nigriventris showing that TH was never co-localized with serotonin. Because TH antibodies are known to reveal mostly or only the TH1 enzyme, we hypothesize that th1-expressing dopamine cells (unlike th2-expressing ones) do not co-localize with serotonin. Since the three investigated species engage in social communication using swim bladder associated musculature, we investigated the serotonergic innervation of the hindbrain vocal or electromotor nuclei initiating the social signal. We found in all three species serotonergic fibers seemingly originating from close-by serotonergic neurons of inferior raphe or anterior spinal cord. Minor differences appear to be rather species-specific than dependent on the type of social communication

Spinal corollary discharge modulates motion sensing during vertebrate locomotion

During active movements, neural replicas of the underlying motor commands may assist in adapting motion-detecting sensory systems to an animal's own behaviour. The transmission of such motor efference copies to the mechanosensory periphery offers a potential predictive substrate for diminishing sensory responsiveness to self-motion during vertebrate locomotion. Here, using semi-isolated in vitro preparations of larval Xenopus, we demonstrate that shared efferent neural pathways to hair cells of vestibular endorgans and lateral line neuromasts express cyclic impulse bursts during swimming that are directly driven by spinal locomotor circuitry. Despite common efferent innervation and discharge patterns, afferent signal encoding at the two mechanosensory peripheries is influenced differentially by efference copy signals, reflecting the different organization of body/water motion-detecting processes in the vestibular and lateral line systems. The resultant overall gain reduction in sensory signal encoding in both cases, which likely prevents overstimulation, constitutes an adjustment to increased stimulus magnitudes during locomotion

Regulation of antioxidant defenses in the prevention of skeletal muscle deconditioning

Musculoskeletal system plays a key role in organism’s well functioning and is responsible for a large variety of functions such as posture, locomotion, balance, and activities of daily life. The quality of the skeletal muscle is therefore capital to maintain quality of life and, in the long term, survival. Hypoactivity and aging are two situations that cause skeletal muscle deconditioning, therefore sharing common characteristics: loss of muscle strength, muscular atrophy and MyHC redistribution, as well as IMAT accumulation. To date, there is plenty of evidence supporting a causative link between oxidative stress phenomenon and muscle deconditioning. The two studies exposed in this thesis demonstrated that modulate antioxidant defense systems have important consequences on deconditioned skeletal muscle tissue. The first study aimed to evaluate frailty in old female animals, using WT and G6PD overexpressing mice. We did a longitudinal functional evaluation, testing the mice each 2 months from 18 to 26 months of age, and then we calculated a frailty score in both groups. In muscle samples of 21-month old mice, we evaluated muscle quality parameters and oxidative stress markers. Finally, we performed a transcriptomic analysis of muscle samples and highlighted differentially expressed genes in both groups of mice. The second study was conducted to evaluate the effects of a cocktail enriched in antioxidant/anti-inflammatory molecules in a 2-month hypoactivity experiment. This countermeasure was expected to limit the effects of muscle deconditioning, but our results clearly demonstrate the ineffectiveness of this type of supplementation in the prevention of muscle mass and strength loss. Moreover, data regarding muscle molecular mechanisms highlight an alteration of recovery processes in the supplemented subjects. These results can be explained by an inhibition of the beneficial adaptations induced by the presence of RONS and illustrate the necessity of pro-oxidant molecules during long-term inactivity to maintain a certain level of muscle function. It underlines the complexity of redox balance mechanisms and demonstrates that physiological amounts of RONS are essential to activate molecular pathways and preserve positive adaptations. Finally, the conclusions of our two studies gave clues on the suitable antioxidant modulation strategy for the prevention of skeletal muscle deconditioning. It seems preferable to focus on the stimulation of endogenous defense system whether than towards exogenous supply of nutritional antioxidants. Nevertheless, the complexity of redox signaling requires better understanding to optimize countermeasures in muscle wasting situations.Musculoskeletal system plays a key role in organism’s well functioning and is responsible for a large variety of functions such as posture, locomotion, balance, and activities of daily life. The quality of the skeletal muscle is therefore capital to maintain quality of life and, in the long term, survival. Hypoactivity and aging are two situations that cause skeletal muscle deconditioning, therefore sharing common characteristics: loss of muscle strength, muscular atrophy and MyHC redistribution, as well as IMAT accumulation. To date, there is plenty of evidence supporting a causative link between oxidative stress phenomenon and muscle deconditioning. The two studies exposed in this thesis demonstrated that modulate antioxidant defense systems have important consequences on deconditioned skeletal muscle tissue. The first study aimed to evaluate frailty in old female animals, using WT and G6PD overexpressing mice. We did a longitudinal functional evaluation, testing the mice each 2 months from 18 to 26 months of age, and then we calculated a frailty score in both groups. In muscle samples of 21-month old mice, we evaluated muscle quality parameters and oxidative stress markers. Finally, we performed a transcriptomic analysis of muscle samples and highlighted differentially expressed genes in both groups of mice. The second study was conducted to evaluate the effects of a cocktail enriched in antioxidant/anti-inflammatory molecules in a 2-month hypoactivity experiment. This countermeasure was expected to limit the effects of muscle deconditioning, but our results clearly demonstrate the ineffectiveness of this type of supplementation in the prevention of muscle mass and strength loss. Moreover, data regarding muscle molecular mechanisms highlight an alteration of recovery processes in the supplemented subjects. These results can be explained by an inhibition of the beneficial adaptations induced by the presence of RONS and illustrate the necessity of pro-oxidant molecules during long-term inactivity to maintain a certain level of muscle function. It underlines the complexity of redox balance mechanisms and demonstrates that physiological amounts of RONS are essential to activate molecular pathways and preserve positive adaptations. Finally, the conclusions of our two studies gave clues on the suitable antioxidant modulation strategy for the prevention of skeletal muscle deconditioning. It seems preferable to focus on the stimulation of endogenous defense system whether than towards exogenous supply of nutritional antioxidants. Nevertheless, the complexity of redox signaling requires better understanding to optimize countermeasures in muscle wasting situation

Measuring flow velocity and flow direction by spatial and temporal analysis of flow fluctuations

If exposed to bulk water flow, fish lateral line afferents respond only to flow fluctuations (AC) and not to the steady (DC) component of the flow. Consequently, a single lateral line afferent can encode neither bulk flow direction nor velocity. It is possible, however, for a fish to obtain bulk flow information using multiple afferents that respond only to flow fluctuations. We show by means of particle image velocimetry that, if a flow contains fluctuations, these fluctuations propagate with the flow. A cross-correlation of water motion measured at an upstream point with that at a downstream point can then provide information about flow velocity and flow direction. In this study, we recorded from pairs of primary lateral line afferents while a fish was exposed to either bulk water flow, or to the water motion caused by a moving object. We confirm that lateral line afferents responded to the flow fluctuations and not to the DC component of the flow, and that responses of many fiber pairs were highly correlated, if they were time-shifted to correct for gross flow velocity and gross flow direction. To prove that a cross-correlation mechanism can be used to retrieve the information about gross flow velocity and direction, we measured the flow-induced bending motions of two flexible micropillars separated in a downstream direction. A cross-correlation of the bending motions of these micropillars did indeed produce an accurate estimate of the velocity vector along the direction of the micropillars

Ontogenetic Development of Vestibulo-Ocular Reflexes in Amphibians

Vestibulo-ocular reflexes (VOR) ensure gaze stability during locomotion and passively induced head/body movements. In precocial vertebrates such as amphibians, vestibular reflexes are required very early at the onset of locomotor activity. While the formation of inner ears and the assembly of sensory-motor pathways is largely completed soon after hatching, angular and translational/tilt VOR display differential functional onsets and mature with different time courses. Otolith-derived eye movements appear immediately after hatching, whereas the appearance and progressive amelioration of semicircular canal-evoked eye movements is delayed and dependent on the acquisition of sufficiently large semicircular canal diameters. Moreover, semicircular canal functionality is also required to tune the initially omnidirectional otolith-derived VOR. The tuning is due to a reinforcement of those vestibulo-ocular connections that are co-activated by semicircular canal and otolith inputs during natural head/body motion. This suggests that molecular mechanisms initially guide the basic ontogenetic wiring, whereas semicircular canal-dependent activity is required to establish the spatio-temporal specificity of the reflex. While a robust VOR is activated during passive head/body movements, locomotor efference copies provide the major source for compensatory eye movements during tail and limb-based swimming of larval and adult frogs. The integration of active/passive motion-related signals for gaze stabilization occurs in central vestibular neurons that are arranged as segmentally iterated functional groups along rhombomere 1-8. However, at variance with the topographic maps of most other sensory systems, the sensory-motor transformation of motion-related signals occurs in segmentally specific neuronal groups defined by the extraocular motor output targets

Intrinsic frequency response patterns in mechano-sensory neurons of the leech

Animals employ mechano-sensory systems to detect and explore their environment. Mechano-sensation encompasses stimuli such as constant pressure, surface movement or vibrations at various intensities that need to be segregated in the central nervous system. Besides different receptor structures, sensory filtering via intrinsic response properties could provide a convenient way to solve this problem. In leech, three major mechano-sensory cell types can be distinguished, according to their stimulus sensitivity, as nociceptive, pressure and touch cells. Using intracellular recordings, we show that the different mechano-sensory neuron classes in Hirudo medicinalis differentially respond supra-threshold to distinct frequencies of sinusoidal current injections between 0.2 and 20 Hz. Nociceptive cells responded with a low-pass filter characteristic, pressure cells as high-pass filters and touch cells as an intermediate band-pass filter. Each class of mechano-sensory neurons is thus intrinsically tuned to a specific frequency range of voltage oscillation that could help segregate mechano-sensory information centrally

Glucose 6-P Dehydrogenase Overexpression Improves Aging-Induced Endothelial Dysfunction in Aorta from Mice: Role of Arginase II

The increase of vascular arginase activity during aging causes endothelial dysfunction. This enzyme competes with the endothelial nitric oxide synthase (eNOS) for L-arginine substrate. Our hypothesis is that glucose 6-P dehydrogenase (G6PD) overexpression could improve the endothelial function modulating the arginase pathway in aorta from mice. For this study, three groups of male mice were used: young wild type (WT) (6-9 months), old WT (21-22 months) and old G6PD-Tg (21-22 months) mice. Vascular reactivity results showed a reduced acetylcholine-dependent relaxation in the old WT but not old G6PD-Tg group. Endothelial dysfunction was reverted by nor-NOHA, an arginase inhibitor. Mice overexpressing G6PD underexpressed arginase II and also displayed a lower activity of this enzyme. Moreover, histological analyses demonstrated that age causes a thickness of aortic walls, but this did not occur in G6PD-Tg mice. We conclude that the overexpressing G6PD mouse is a model to improve vascular health via the arginase pathway