Light-stimulus intensity modulates startle reflex habituation in larval zebrafish.

The startle reflex in larval zebrafish describes a C-bend of the body occurring in response to sudden, unexpected, stimuli of different sensory modalities. Alterations in the startle reflex habituation (SRH) have been reported in various human and animal models of neurological and psychiatric conditions and are hence considered an important behavioural marker of neurophysiological function. The amplitude, offset and decay constant of the auditory SRH in larval zebrafish have recently been characterised, revealing that the measures are affected by variation in vibratory frequency, intensity, and interstimulus-interval. Currently, no study provides a model-based analysis of the effect of physical properties of light stimuli on the visual SRH. This study assessed the effect of incremental light-stimulus intensity on the SRH of larval zebrafish through a repeated-measures design. Their total locomotor responses were normalised for the time factor, based on the behaviour of a (non-stimulated) control group. A linear regression indicated that light intensity positively predicts locomotor responses due to larger SRH decay constants and offsets. The conclusions of this study provide important insights as to the effect of light properties on the SRH in larval zebrafish. Our methodology and findings constitute a relevant reference framework for further investigation in translational neurophysiological research.

Dual Color Neural Activation and Behavior Control with Chrimson and CoChR in Caenorhabditis elegans.

By enabling a tight control of cell excitation, optogenetics is a powerful approach to study the function of neurons and neural circuits. With its transparent body, a fully mapped nervous system, easily quantifiable behaviors and many available genetic tools, Caenorhabditis elegans is an extremely well-suited model to decipher the functioning logic of the nervous system with optogenetics. Our goal was to establish an efficient dual color optogenetic system for the independent excitation of different neurons in C. elegans. We combined two recently discovered channelrhodopsins: the red-light sensitive Chrimson from Chlamydomonas noctigama and the blue-light sensitive CoChR from Chloromonas oogama. Codon-optimized versions of Chrimson and CoChR were designed for C. elegans and expressed in different mechanosensory neurons. Freely moving animals produced robust behavioral responses to light stimuli of specific wavelengths. Since CoChR was five times more sensitive to blue light than the commonly used ChR2, we were able to use low blue light intensities producing no cross-activation of Chrimson. Thanks to these optogenetics tools, we revealed asymmetric cross-habituation effects between the gentle and harsh touch sensory motor pathways. Collectively, our results establish the Chrimson/CoChR pair as a potent tool for bimodal neural excitation in C. elegans and equip this genetic model organism for the next generation of in vivo optogenetic analyses.

Habituation of steady-state visual evoked potentials in response to high-frequency polychromatic foveal visual stimulation.

In an attempt to develop safe and robust methods for monitoring migraineurs' brain states, we explores the feasibility of using white, red, green and blue LED lights flickering around their critical flicker fusion (CFF) frequencies as foveal visual stimuli for inducing steady-state visual evoked potentials (SSVEP) and causing discernible habituation trends. After comparing the habituation indices, the multi-scale entropies and the time dependent intrinsic correlations of their SSVEP signals, we reached a tentative conclusion that sharp red and white light pulses flickering barely above their CFF frequencies can replace commonly used 13Hz stimuli to effectively cause SSVEP habituation among normal subjects. Empirical results showed that consecutive short bursts of light can produce more consistent responses than a single prolonged stimulation. Since these high frequency stimuli do not run the risk of triggering migraine or seizure attacks, further tests of these stimuli on migraine patients are warranted in order to verify their effectiveness.

Photoperiod mediated changes in olfactory bulb neurogenesis and olfactory behavior in male white-footed mice (Peromyscus leucopus).

Brain plasticity, in relation to new adult mammalian neurons generated in the subgranular zone of the hippocampus, has been well described. However, the functional outcome of new adult olfactory neurons born in the subventricular zone of the lateral ventricles is not clearly defined, as manipulating neurogenesis through various methods has given inconsistent and conflicting results in lab mice. Several small rodent species, including Peromyscus leucopus, display seasonal (photoperiodic) brain plasticity in brain volume, hippocampal function, and hippocampus-dependent behaviors; plasticity in the olfactory system of photoperiodic rodents remains largely uninvestigated. We exposed adult male P. leucopus to long day lengths (LD) and short day lengths (SD) for 10 to 15 weeks and then examined olfactory bulb cell proliferation and survival using the thymidine analog BrdU, olfactory bulb granule cell morphology using Golgi-Cox staining, and behavioral investigation of same-sex conspecific urine. SD mice did not differ from LD counterparts in granular cell morphology of the dendrites or in dendritic spine density. Although there were no differences due to photoperiod in habituation to water odor, SD mice rapidly habituated to male urine, whereas LD mice did not. In addition, short day induced changes in olfactory behavior were associated with increased neurogenesis in the caudal plexiform and granule cell layers of the olfactory bulb, an area known to preferentially respond to water-soluble odorants. Taken together, these data demonstrate that photoperiod, without altering olfactory bulb neuronal morphology, alters olfactory bulb neurogenesis and olfactory behavior in Peromyscus leucopus.

Investigation of the effects of head irradiation with gamma rays and protons on startle and pre-pulse inhibition behavior in mice.

With the increased international emphasis on manned space exploration, there is a growing need to understand the impact of the spaceflight environment on health and behavior. One particularly important aspect of this environment is low-dose radiation. In the present studies, we first characterized the γ- and proton-irradiation dose effect on acoustic startle and pre-pulse inhibition behaviors in mice exposed to 0-5 Gy brain-localized irradiation, and assessed these effects 2 days later. Subsequently, we used 2 Gy to assess the time course of γ- and proton-radiation effects on startle reactivity 0-8 days after exposure. Exposures targeted the brain to minimize the impact of peripheral inflammation-induced sickness behavior. The effects of radiation on startle were subtle and acute. Radiation reduced the startle response at 2 and 5 Gy. Following a 2-Gy exposure, the response reached a minimum at the 2-day point. Proton and γ-ray exposures did not differ in their impact on startle. We found there were no effects of radiation on pre-pulse inhibition of the startle response.

An early treatment with 17-ß-estradiol is neuroprotective against the long-term effects of neonatal ionizing radiation exposure.

Ionizing radiations can induce oxidative stress on target tissues, acting mainly through reactive oxygen species (ROS). The aim of this work was to investigate if 17-ß-estradiol (ßE) was able to prevent hippocampal-related behavioral and biochemical changes induced by neonatal ionizing radiation exposure and to elucidate a potential neuroprotective mechanism. Male Wistar rats were irradiated with 5 Gy of X-rays between 24 and 48 h after birth. A subset of rats was subcutaneously administered with successive injections of ßE or 17-α-estradiol (αE), prior and after irradiation. Rats were subjected to different behavioral tasks to evaluate habituation and associative memory as well as anxiety levels. Hippocampal ROS levels and protein kinase C (PKC) activity were also assessed. Results show that although ßE was unable to prevent radiation-induced hippocampal PKC activity changes, most behavioral abnormalities were reversed. Moreover, hippocampal ROS levels in ßE-treated irradiated rats approached control values. In addition, αE administered to irradiated animals was effective in preventing radiation-induced alterations. In conclusion, ßE was able to counteract behavioral and biochemical changes induced in irradiated animals, probably acting through an antioxidant mechanism.

Pain perception and laser evoked potentials during menstrual cycle in migraine.

The association between estrogens "withdrawal" and attacks of migraine without aura is well-known. The aim of the study was to examine the features of laser evoked potentials (LEPs), including habituation, in women suffering from migraine without aura versus healthy controls, during the pre-menstrual and late luteal phases. Nine migraine without aura and 10 non-migraine healthy women, were evaluated during the pre-menstrual phase and late luteal phase. The LEPs were recorded during the inter-critical phase. The right supraorbital zone and the dorsum of the right hand were stimulated. Three consecutive series of 20 laser stimuli were obtained for each stimulation site. Laser pain perception was rated by a 0-100 VAS after each stimulation series. Migraine patients exhibited increased LEPs amplitude and reduced habituation compared to normal subjects. Laser-pain perception was increased during the pre-menstrual phase in both patients and controls. Migraine patients and controls showed increased P2 and N2-P2 amplitude in the pre-menstrual phase, on both stimulation sites. During the pre-menstrual phase the N2-P2 habituation appeared to be reduced in both migraine and healthy women. The estrogen withdrawal occurring during the menstrual cycle may favor reduced habituation of nociceptive cortex, which may facilitate pain symptoms and migraine in predisposed women.

Mobile phones exposure induces changes of contingent negative variation in humans.

Event-related potentials have been largely employed to test effects of GSM emissions on human brain. The aim of the present study was the evaluation of initial contingent negative variation (iCNV) changes, induced by 900 MHz GSM exposure, in a double blind design in healthy volunteers, subjected to a threefold experimental condition, EXPOSED (A), a real GSM phone emitting electromagnetic power, SHAM (B), a real phone where the electromagnetic power was dissipated on an internal load and OFF (C), a phone completely switched-off. Ten healthy right-handed volunteers were evaluated. The CNV was recorded during a 10 min time interval in each of the three experimental conditions A, B, and C, in order to assess the iCNV amplitude and habituation. The iCNV amplitude decreased and habituation increased during both A and B conditions, compared with condition C. This effect was diffuse over the scalp, and there was no significant prevalence of iCNV amplitude reduction on the left side, were the phones were located. Mobile Phones exposures A and B seemed to act on brain electrical activity, reducing the arousal and expectation of warning stimulus. This evidence, limited by the low number of subjects investigated, could be explained in terms of an effect induced by both the GSM signal and the extremely low frequency magnetic field produced by battery and internal circuits.

Paradoxical shortening of sympathetic skin response latency at distal recording sites.

OBJECTIVE: The purpose of this study was to investigate and to discuss the neurophysiological mechanism of paradoxical shortening of the sympathetic skin response (SSR) latency at distal recording sites. METHODS: The latency and peak-to-peak amplitude of SSRs evoked by magnetic stimuli were analyzed. Eight active electrodes were placed on the palmar (anterior) and dorsal (posterior) sides of the hand (forearm) proximal to the distal arrangement. RESULTS: SSRs from two palm sites had significantly shorter latencies and larger amplitudes than the SSRs at the other six sites, including the proximal sites of the forearm. CONCLUSIONS: This finding indicated that the SSR latency at different sites was not linearly prolonged as the distance of the recording sites from the proximal to distal areas increased. The paradoxical shortening of the latency and the large amplitude of the SSR from the palm can be explained by a recent model of the equivalent current dipole caused by the Na+ concentration gradient. The high density of sweat glands in the palm possibly produced the present findings. SIGNIFICANCE: We should carefully interpret the sudomotor conduction velocity derived from latency difference between two sites, especially for thermal and emotional sweating.

Extremely low-frequency magnetic fields effects on the snail single neurons.

The aim of present work is to explore the influence of extremely low-frequency electromagnetic fields (8.34 and 217 Hz) utilized in cell phones on habituation of the mollusk single neuron to intracellular stimuli. The isolated nervous system of the mollusk Helix Pomatia was used in the experiments. Helmholtz coils were used to expose brain ganglia to the low-frequency electromagnetic fields. Peak values of the extremely low-frequency fields were between 1 and 6 mT. Neuron electrophysiology was investigated using a standard microelectrode technique. Exposure of the neuron to the low-frequency electromagnetic fields caused dehabituation to intracellular stimulus. The effect was proportional to the magnetic induction peak value. The observed dehabituation occurs by degradation of the signal to noise ratio and by alteration of the neuron's normal function.

Open field behavior and habituation in rats irradiated on the head with gamma-rays.

The goal of this work was to establish the effects of irradiation of the head with gamma-rays on selected forms of innate behavior, as well as on the habituation process to new environment in the open field test in rats. During a 5-day control period behavioral parameters reflecting motoric and explorative activities, as well as anxiety (6 parameters in total) were followed daily in 20 male Sprague-Dawley rats in an open field. Fourteen animals were repeatedly tested after irradiation of the head with a single dose of 10 Gy of gamma-rays. The results showed statistically significant depression of motoric and explorative activities during the first 3 days after irradiation in comparison with the control period. After irradiation the level of anxiety was elevated. Habituation to a new environment was observed in non-irradiated, but not in irradiated animals. These results suggest that the applied dose of ionizing radiation influenced the brain centers involved in control of innate behavioral functions connected with response to a new environment in rats.

Ischemia-induced hyperactivity: effects of dim versus bright illumination on open-field exploration and habituation following global ischemia in rats.

The current study reports the impact of different illumination conditions on exploratory activity following global ischemia in rats. Exploratory activity was tested at different post-ischemic intervals under bright (450 lux) or dim (40 lux) light exposure. A 30 min testing period performed 5 days post-reperfusion examined within-session open-field habituation in ischemic and sham-operated animals. Additional animals were tested in the open-field under the two illumination conditions for shorter 10 min tests on days 3, 6, and 9 following reperfusion. Our findings demonstrated illumination-related activity profile in the open-field in ischemic animals. While ischemic rats showed increased activity when tested under bright open-field illumination, reduced activity was observed under dim illumination as compared to sham-operated controls. Further, habituation deficits were not apparent in animals subjected to global ischemia under any illumination condition. Similar behavioral profiles and habituation were observed in ischemic animals when exposed to repetitive open-field tests at days 3, 6, and 9 following reperfusion. CA1 neuronal injury (approximately 75% as compared to sham rats) was comparable in all ischemic groups at day 12 following reperfusion. The present findings suggest that differences in initial behavioral reactivity of sham and ischemic rats to bright versus dimly lighted environments may contribute to differences in open-field exploration reported between these groups. They also challenge the notion that deficits in exploration in ischemic animals are mainly attributable to processes related to habituation, or that hyperactivity represents a reliable predictor of CA1 neuronal injury. These observations may help explain discrepant ischemia-induced behavioral effects reported in the open field.

Hippocampal neurogenesis is not required for behavioral effects of environmental enrichment.

Environmental enrichment increases adult hippocampal neurogenesis and alters hippocampal-dependent behavior in rodents. To investigate a causal link between these two observations, we analyzed the effect of enrichment on spatial learning and anxiety-like behavior while blocking adult hippocampal neurogenesis. We report that environmental enrichment alters behavior in mice regardless of their hippocampal neurogenic capability, providing evidence that the newborn cells do not mediate these effects of enrichment.

Synaptic depression and short-term habituation are located in the sensory part of the mammalian startle pathway.

BACKGROUND: Short-term habituation of the startle response represents an elementary form of learning in mammals. The underlying mechanism is located within the primary startle pathway, presumably at sensory synapses on giant neurons in the caudal pontine reticular nucleus (PnC). Short trains of action potentials in sensory afferent fibers induce depression of synaptic responses in PnC giant neurons, a phenomenon that has been proposed to be the cellular correlate for short-term habituation. We address here the question whether both this synaptic depression and the short-term habituation of the startle response are localized at the presynaptic terminals of sensory afferents. If this is confirmed, it would imply that these processes take place prior to multimodal signal integration, rather than occurring at postsynaptic sites on PnC giant neurons that directly drive motor neurons. RESULTS: Patch-clamp recordings in vitro were combined with behavioral experiments; synaptic depression was specific for the input pathway stimulated and did not affect signals elicited by other sensory afferents. Concordant with this, short-term habituation of the acoustic startle response in behavioral experiments did not influence tactile startle response amplitudes and vice versa. Further electrophysiological analysis showed that the passive properties of the postsynaptic neuron were unchanged but revealed some alterations in short-term plasticity during depression. Moreover, depression was induced only by trains of presynaptic action potentials and not by single pulses. There was no evidence for transmitter receptor desensitization. In summary, the data indicates that the synaptic depression mechanism is located presynaptically. CONCLUSION: Our electrophysiological and behavioral data strongly indicate that synaptic depression in the PnC as well as short-term habituation are located in the sensory part of the startle pathway, namely at the axon terminals of sensory afferents in the PnC. Our results further corroborate the link between synaptic depression and short-term habituation of the startle response.

Sensitization and habituation of AMH and C-fiber related percepts of repetitive radiant heat stimulation.

OBJECTIVE: Pain perception involves neuronal plasticity at peripheral and central stages, resulting in sensitization or habituation, depending on intensity and temporal features of stimulation. Concurrent assessment of perceptual change over different time spans is therefore important for understanding the dynamics of pain processing. METHODS: A new psychophysical procedure was established to assess sensitization and habituation during repetitive radiant heat stimulation. Short-term perceptual change (<1 min) during trials with 10 stimuli applied at 3 frequencies (0.2-0.6-1.8 Hz) and 3 intensities was assessed for AMH-II or C-fiber related percepts. Perceptual changes were monitored for medium-term (1-15 min) and for long-term (15-90 min) time spans. RESULTS: Short-term sensitization occurred only at frequencies above 0.3 Hz and was affected by both stimulus frequency and intensity, but the AMH-fiber related sensitization depended on intensity only above 0.6 Hz. Multiple stimulation of the same skin area during medium-term time spans resulted in habituation. No long-term perceptual changes occurred. CONCLUSIONS: The procedure permits concurrent assessment of short-term sensitization and medium-term habituation, assumed to be related to spinal windup and cutaneous nociceptive fiber fatigue, respectively. SIGNIFICANCE: The method is suitable for quantitative sensory testing of dynamic pain processing over different time spans, relevant in clinical testing of pain and in drug assessment.

Drosophila CAKI/CMG protein, a homolog of human CASK, is essential for regulation of neurotransmitter vesicle release.

Vertebrate CASK is a member of the membrane-associated guanylate kinase (MAGUK) family of proteins. CASK is present in the nervous system where it binds to neurexin, a transmembrane protein localized in the presynaptic membrane. The Drosophila homologue of CASK is CAKI or CAMGUK. CAKI is expressed in the nervous system of larvae and adult flies. In adult flies, the expression of caki is particularly evident in the visual brain regions. To elucidate the functional role of CASK, we employed a caki null mutant in the model organism Drosophila melanogaster. By means of electrophysiological methods, we analyzed, in adult flies, the spontaneous and evoked neurotransmitter release at the neuromuscular junction (NMJ) as well as the functional status of the giant fiber pathway and of the visual system. We found that in caki mutants, when synaptic activity is modified, the spontaneous neurotransmitter release of the indirect flight muscle NMJ was increased, the response of the giant fiber pathway to continuous stimulation was impaired, and electroretinographic responses to single and continuous repetitive stimuli were altered and optomotor behavior was abnormal. These results support the involvement of CAKI in neurotransmitter release and nervous system function.

Validation of an auditory startle response system using chemicals or parametric modulation as positive controls.

Neurotoxicity regulatory guidelines mandate that automated test systems be validated using chemicals. However, in some cases, chemicals may not necessarily be needed to prove test system validity. To examine this issue, two independent experiments were conducted to validate an automated auditory startle response (ASR) system. In Experiment 1, we used adult (PND 63) and weanling (PND 22) Sprague-Dawley rats (10/sex/dose) to determine the effect of either d-amphetamine (4.0 or 8.0 mg/kg) or clonidine (0.4 or 0.8 mg/kg) on the ASR peak amplitude (ASR PA). The startle response of each rat to a short burst of white noise (120 dB SPL) was recorded over 50 consecutive trials. The ASR PA was significantly decreased (by clonidine) and increased (by d-amphetamine) compared to controls in PND 63 rats. In PND 22 rats, the response to clonidine was similar to adults, but d-amphetamine effects were not significant. Neither drug affected the rate of the decrease in ASR PA over time (habituation). In Experiment 2, PND 31 Sprague-Dawley rats (8/sex) were presented with 150 trials consisting of either white noise bursts of variable intensity (70-120 dB SPL in 10 dB increments, presented in random order) or null (0 dB SPL) trials. Statistically significant sex- and intensity-dependent differences were detected in the ASR PA. These results suggest that in some cases, parametric modulation may be an alternative to using chemicals for test system validation.

Effects of exposure to extremely low-frequency magnetic field of 2 G intensity on memory and corticosterone level in rats.

In the present study, we examined the effects of chronic exposure (1 and 2 weeks) to an extremely low-frequency magnetic field (ELFMF) of 2 G intensity on memory in rats using an object recognition task. Comparable groups of rats were exposed for 1, 2 or 4 weeks to ELFMF and the following day blood samples were collected from each rat for the measurement of corticosterone level. Our results demonstrate that exposure to ELFMF induces a significant increase in the level of corticosterone in blood plasma and is associated with impairment in discrimination between familiar and novel objects.