Wide Range Of Stimulus Intensities Research Articles

Diversity matters - extending sound intensity coding by inner hair cells via heterogeneous synapses.

Our sense of hearing enables the processing of stimuli that differ in sound pressure by more than six orders of magnitude. How to process a wide range of stimulus intensities with temporal precision is an enigmatic phenomenon of the auditory system. Downstream of dynamic range compression by active cochlear micromechanics, the inner hair cells (IHCs) cover the full intensity range of sound input. Yet, the firing rate in each of their postsynaptic spiral ganglion neurons (SGNs) encodes only a fraction of it. As a population, spiral ganglion neurons with their respective individual coding fractions cover the entire audible range. How such "dynamic range fractionation" arises is a topic of current research and the focus of this review. Here, we discuss mechanisms for generating the diverse functional properties of SGNs and formulate testable hypotheses. We postulate that an interplay of synaptic heterogeneity, molecularly distinct subtypes of SGNs, and efferent modulation serves the neural decomposition of sound information and thus contributes to a population code for sound intensity.

Stimulus Driven Functional Transformations in the Early Olfactory System.

Olfactory stimuli are encountered across a wide range of odor concentrations in natural environments. Defining the neural computations that support concentration invariant odor perception, odor discrimination, and odor-background segmentation across a wide range of stimulus intensities remains an open question in the field. In principle, adaptation could allow the olfactory system to adjust sensory representations to the current stimulus conditions, a well-known process in other sensory systems. However, surprisingly little is known about how adaptation changes olfactory representations and affects perception. Here we review the current understanding of how adaptation impacts processing in the first two stages of the vertebrate olfactory system, olfactory receptor neurons (ORNs), and mitral/tufted cells.

Exocytosis in mouse vestibular Type II hair cells shows a high-order Ca2+ dependence that is independent of synaptotagmin-4.

Mature hair cells transduce information over a wide range of stimulus intensities and frequencies for prolonged periods of time. The efficiency of such a demanding task is reflected in the characteristics of exocytosis at their specialized presynaptic ribbons. Ribbons are electron‐dense structures able to tether a large number of releasable vesicles allowing them to maintain high rates of vesicle release. Calcium entry through rapidly activating, non‐inactivating CaV1.3 (L‐type) Ca2+ channels in response to cell depolarization causes a local increase in Ca2+ at the ribbon synapses, which is detected by the exocytotic Ca2+ sensors. The Ca2+ dependence of vesicle exocytosis at mammalian vestibular hair cell (VHC) ribbon synapses is believed to be linear, similar to that observed in mature cochlear inner hair cells (IHCs). The linear relation has been shown to correlate with the presence of the Ca2+ sensor synaptotagmin‐4 (Syt‐4). Therefore, we studied the exocytotic Ca2+ dependence, and the release kinetics of different vesicle pool populations, in Type II VHCs of control and Syt‐4 knockout mice using patch‐clamp capacitance measurements, under physiological recording conditions. We found that exocytosis in mature control and knockout Type II VHCs displayed a high‐order dependence on Ca2+ entry, rather than the linear relation previously observed. Consistent with this finding, the Ca2+ dependence and release kinetics of the ready releasable pool (RRP) of vesicles were not affected by an absence of Syt‐4. However, we did find that Syt‐4 could play a role in regulating the release of the secondary releasable pool (SRP) in these cells. Our findings show that the coupling between Ca2+ influx and neurotransmitter release at mature Type II VHC ribbon synapses is faithfully described by a nonlinear relation that is likely to be more appropriate for the accurate encoding of low‐frequency vestibular information, consistent with that observed at low‐frequency mammalian auditory receptors.

What range of stimulus intensities should we apply to elicit abnormal muscle response in microvascular decompression for hemifacial spasm?

Abnormal muscle response (AMR) has been considered as a predictor of the prognosis after microvascular decompression (MVD) for hemifacial spasm (HFS). However, its predictive value has not always been satisfactory. The objective of this work was to confirm an optimal range of stimulus intensities to elicit AMR in surgery. Seventy-two consecutive patients with primary HFS treated by MVD were retrospectively included in this study. A wide range of stimulus intensities from 1 to 100mA was applied in AMR monitoring. The AMR-elicited threshold value was quantitatively traced throughout all surgical procedures. The relationship between clinical outcomes and electrophysiological findings was analyzed. Of the 72 patients, 44 were immediately cured and 24 were delayed cured; the remaining 4 were proved not to be cured in their follow-up periods. The patterns of AMR-elicited threshold changes were categorized into five types, which could only be discriminated with a wide range of stimulus intensities. The constituent ratio of the patterns was significantly different (P < 0.001) among the clinical outcomes. Some patterns of AMR changes might have been ignored if we had only applied a narrow range of stimulus intensities (1-30mA) to judge whether AMR disappeared or not. Thus, a wide range of stimulus intensities (1-100mA) to trace the AMR-elicited threshold values was proposed for a more precise prediction.

Characterization of physiological phenotypes of dentate gyrus synapses of PDZ1/2 domain-deficient PSD-95-knockin mice.

The hippocampal formation is involved in several important brain functions of animals, such as memory formation and pattern separation, and the synapses in the dentate gyrus (DG) play critical roles as the first step in the hippocampal circuit. Previous studies have reported that mice with genetic modifications of the PDZ1/2 domains of postsynaptic density (PSD)-95 exhibit altered synaptic properties in the DG and impaired hippocampus-dependent behaviors. Based on the involvement of the DG in the regulation of behaviors, these data suggest that the abnormal behavior of these knockin (KI) mice is due partly to altered DG function. Precise understanding of the phenotypes of these mutant mice requires characterization of the synaptic properties of the DG, and here we provide detailed studies of DG synapses. We have demonstrated global changes in the PSD membrane-associated guanylate kinase expression pattern in the DG of mutant mice, and DG synapses in these mice exhibited increased long-term potentiation under a wide range of stimulus intensities, although the N-methyl-d-aspartic acid receptor dependence of the long-term potentiation was unchanged. Furthermore, our data also indicate increased silent synapses in the DG of the KI mice. These findings suggest that abnormal protein expression and physiological properties disrupt the function of DG neurons in these KI mice.

Faithful Representation of Tactile Intensity under Different Contexts Emerges from the Distinct Adaptive Properties of the First Somatosensory Relay Stations.

Adaptation allows neurons to respond to a wide range of stimulus intensities. However, it also leads to ambiguity as the representation of the external world depends on the context. We recorded neurons from Wistar rats' brainstem nuclei belonging to two major somatosensory pathways (lemniscal and paralemniscal) and explored the way in which they encode noisy stimuli under different contexts. We found that although their unadapted intensity-response curves are very similar, the adapted curves of the two pathways are distinctively different as they are optimized for encoding different intensity ranges. Lemniscal neurons most faithfully encoded stimuli when the background intensity was high, whereas paralemniscal cells best encoded stimuli under low intensity context. Intracellular recordings indicate that these differences emerge already at the synaptic level. We suggest that the two pathways synergistically improve the ability of this system to encode a wide range of intensities during natural stimulation, potentially reducing the inherent ambiguity of adaptive coding.

Circuit motifs for contrast-adaptive differentiation in early sensory systems: the role of presynaptic inhibition and short-term plasticity.

In natural signals, such as the luminance value across of a visual scene, abrupt changes in intensity value are often more relevant to an organism than intensity values at other positions and times. Thus to reduce redundancy, sensory systems are specialized to detect the times and amplitudes of informative abrupt changes in the input stream rather than coding the intensity values at all times. In theory, a system that responds transiently to fast changes is called a differentiator. In principle, several different neural circuit mechanisms exist that are capable of responding transiently to abrupt input changes. However, it is unclear which circuit would be best suited for early sensory systems, where the dynamic range of the natural input signals can be very wide. We here compare the properties of different simple neural circuit motifs for implementing signal differentiation. We found that a circuit motif based on presynaptic inhibition (PI) is unique in a sense that the vesicle resources in the presynaptic site can be stably maintained over a wide range of stimulus intensities, making PI a biophysically plausible mechanism to implement a differentiator with a very wide dynamical range. Moreover, by additionally considering short-term plasticity (STP), differentiation becomes contrast adaptive in the PI-circuit but not in other potential neural circuit motifs. Numerical simulations show that the behavior of the adaptive PI-circuit is consistent with experimental observations suggesting that adaptive presynaptic inhibition might be a good candidate neural mechanism to achieve differentiation in early sensory systems.

Light adaptation alters the source of inhibition to the mouse retinal OFF pathway

Sensory systems must avoid saturation to encode a wide range of stimulus intensities. One way the retina accomplishes this is by using both dim-light-sensing rod and bright-light-sensing cone photoreceptor circuits. OFF cone bipolar cells are a key point in this process, as they receive both excitatory input from cones and inhibitory input from AII amacrine cells via the rod pathway. However, in addition to AII amacrine cell input, other inhibitory inputs from cone pathways also modulate OFF cone bipolar cell light signals. It is unknown how these inhibitory inputs to OFF cone bipolar cells change when switching between rod and cone pathways or whether all OFF cone bipolar cells receive rod pathway input. We found that one group of OFF cone bipolar cells (types 1, 2, and 4) receive rod-mediated inhibitory inputs that likely come from the rod-AII amacrine cell pathway, while another group of OFF cone bipolar cells (type 3) do not. In both cases, dark-adapted rod-dominant light responses showed a significant contribution of glycinergic inhibition, which decreased with light adaptation and was, surprisingly, compensated by an increase in GABAergic inhibition. As GABAergic input has distinct timing and spatial spread from glycinergic input, a shift from glycinergic to GABAergic inhibition could significantly alter OFF cone bipolar cell signaling to downstream OFF ganglion cells. Larger GABAergic input could reflect an adjustment of OFF bipolar cell spatial inhibition, which may be one mechanism that contributes to retinal spatial sensitivity in the light.

Transmitter Release from Cochlear Hair Cells Is Phase Locked to Cyclic Stimuli of Different Intensities and Frequencies

The auditory system processes time and intensity through separate brainstem pathways to derive spatial location as well as other salient features of sound. The independent coding of time and intensity begins in the cochlea, where afferent neurons can fire action potentials at constant phase throughout a wide range of stimulus intensities. We have investigated time and intensity coding by simultaneous presynaptic and postsynaptic recording at the hair cell-afferent synapse from rats. Trains of depolarizing steps to the hair cell were used to elicit postsynaptic currents that occurred at constant phase for a range of membrane potentials over which release probability varied significantly. To probe the underlying mechanisms, release was examined using single steps to various command voltages. As expected for vesicular release, first synaptic events occurred earlier as presynaptic calcium influx grew larger. However, synaptic depression produced smaller responses with longer first latencies. Thus, during repetitive hair cell stimulation, as the hair cell is more strongly depolarized, increased calcium channel gating hurries transmitter release, but the resulting vesicular depletion produces a compensatory slowing. Quantitative simulation of ribbon function shows that these two factors varied reciprocally with hair cell depolarization (stimulus intensity) to produce constant synaptic phase. Finally, we propose that the observed rapid vesicle replenishment would help maintain the vesicle pool, which in turn would equilibrate with the stimulus intensity (and therefore the number of open Ca(2+) channels), so that for trains of different levels the average phase will be conserved.

Coding of sound direction in the auditory periphery of the lake sturgeon,Acipenser fulvescens

The lake sturgeon, Acipenser fulvescens, belongs to one of the few extant nonteleost ray-finned fishes and diverged from the main vertebrate lineage about 250 million years ago. The aim of this study was to use this species to explore the peripheral neural coding strategies for sound direction and compare these results to modern bony fishes (teleosts). Extracellular recordings were made from afferent neurons innervating the saccule and lagena of the inner ear while the fish was stimulated using a shaker system. Afferents were highly directional and strongly phase locked to the stimulus. Directional response profiles resembled cosine functions, and directional preferences occurred at a wide range of stimulus intensities (spanning at least 60 dB re 1 nm displacement). Seventy-six percent of afferents were directionally selective for stimuli in the vertical plane near 90° (up down) and did not respond to horizontal stimulation. Sixty-two percent of afferents responsive to horizontal stimulation had their best axis in azimuths near 0° (front back). These findings suggest that in the lake sturgeon, in contrast to teleosts, the saccule and lagena may convey more limited information about the direction of a sound source, raising the possibility that this species uses a different mechanism for localizing sound. For azimuth, a mechanism could involve the utricle or perhaps the computation of arrival time differences. For elevation, behavioral strategies such as directing the head to maximize input to the area of best sensitivity may be used. Alternatively, the lake sturgeon may have a more limited ability for sound source localization compared with teleosts.

Dose-dependent opposite effects of gabapentin on the depressive action of morphine on a C-fibre reflex in the rat

Gabapentin is a structural analogue of gamma-amino-butyric acid with anticonvulsant activity. Recently, indications for its use were extended to the management of acute pain in the postoperative period. The effects of pre-administration of gabapentin on the depressive action of intravenous morphine were studied on the C-fibre reflex elicited by a wide range of stimulus intensities. The reflex was elicited by electrical stimulation of the sural nerve and recorded from the ipsilateral biceps femoris muscle in halothane anaesthetized rats with either an intact neuraxis or a brainstem previously transected at the level of the obex. As previously reported, 6 mg/kg intravenous morphine both increased the threshold and decreased the slope of the stimulus–response recruitment curve. The C-fibre reflex was not modified following intravenous gabapentin. Gabapentin pre-treatment at lower doses (0.01–7.5 mg/kg) not only antagonized the depressive effect of morphine, but caused facilitation of the reflex. At higher doses (10–50 mg/kg), gabapentin pre-treatment potentiated the depressive effect of morphine. In obex-transected rats, the facilitation of the C-fibre reflex, seen following 1 mg/kg gabapentin and 6 mg/kg morphine, disappeared and was replaced by a strong reinforcement of the depressive effect of morphine. It is concluded that a strong synergy between the effects of gabapentin and morphine can be seen at the spinal level. However, radically opposite effects with supraspinal origins thwart this mechanism. From the clinical standpoint, these results incite cautiousness in the use of combinations of gabapentin and opioids.

Enhanced excitability of the corticospinal pathway of the ankle extensor and flexor muscles during standing in humans

The purpose of this study was to investigate how the recruitment properties of the corticospinal pathway are modulated in the soleus (SOL) and tibialis anterior (TA) muscles depending on postures. A wide range of stimulus intensities were applied via transcranial magnetic stimulation over the primary motor cortex during standing (STD) and sitting (SIT) with a comparable background activity level in each muscle. The relationship between the stimulation intensities and the size of motor-evoked potentials was assessed by the Boltzmann sigmoid function, which is characterized by a plateau value, maximum slope, and threshold. The plateau value and maximum slope were significantly higher during STD than during SIT in the SOL muscle (STD vs. SIT, plateau value: 50.0 +/- 21.8 vs. 33.9 +/- 12.3 mV ms, maximal slope: 1.6 +/- 0.7 vs. 1.2 +/- 0.5 mV ms/% maximal stimulator output). Similar changes of the parameters were also observed in the TA muscle (STD vs. SIT, plateau value: 71.0 +/- 22.9 vs. 41.4 +/- 16.1 mV ms, maximal slope: 5.0 +/- 2.0 vs. 2.5 +/- 0.7 mV ms/% maximal stimulator output). The threshold did not differ significantly between the two conditions and both muscles. These results indicate that the central nervous system requires a different control for each postural condition; that is, the relative balance of the excitatory and inhibitory inputs to the corticospinal pathways as well as the number of neurons of subliminal fringe in the corticospinal pathway was increased during STD compared with those during SIT.

Stimulus-Response Profile during Single-Pulse Transcranial Magnetic Stimulation to the Primary Motor Cortex

We examined the stimulus-response profile during single-pulse transcranial magnetic stimulation (TMS) by measuring motor-evoked potentials (MEPs) with electromyographic monitoring and hemodynamic responses with functional magnetic resonance imaging (fMRI) at 3 Tesla. In 16 healthy subjects, single TMS pulses were irregularly delivered to the left primary motor cortex at a mean frequency of 0.15 Hz with a wide range of stimulus intensities. The measurement of MEP proved a typical relationship between stimulus intensity and MEP amplitude in the concurrent TMS-fMRI environment. In the population-level analysis of the suprathreshold stimulation conditions, significant increases in hemodynamic responses were detected in the motor/somatosensory network, reflecting both direct and remote effects of TMS, and also the auditory/cognitive areas, perhaps related to detection of clicks. The stimulus-response profile showed both linear and nonlinear components in the direct and remote motor/somatosensory network. A detailed analysis suggested that the nonlinear components of the motor/somatosensory network activity might be induced by nonlinear recruitment of neurons in addition to sensory afferents resulting from movement. These findings expand our basic knowledge of the quantitative relationship between TMS-induced neural activations and hemodynamic signals measured by neuroimaging techniques.

Evidence for functional alterations in the skeletal muscle mechanoreflex and metaboreflex in hypertensive rats

Exercise in hypertensive individuals elicits exaggerated increases in mean arterial pressure (MAP) and heart rate (HR) that potentially enhance the risk for adverse cardiac events or stroke. Evidence suggests that exercise pressor reflex function (EPR; a reflex originating in skeletal muscle) is exaggerated in this disease and contributes significantly to the potentiated cardiovascular responsiveness. However, the mechanism of EPR overactivity in hypertension remains unclear. EPR function is mediated by the muscle mechanoreflex (activated by stimulation of mechanically sensitive afferent fibers) and metaboreflex (activated by stimulation of chemically sensitive afferent fibers). Therefore, we hypothesized the enhanced cardiovascular response mediated by the EPR in hypertension is due to functional alterations in the muscle mechanoreflex and metaboreflex. To test this hypothesis, mechanically and chemically sensitive afferent fibers were selectively activated in normotensive Wistar-Kyoto (WKY) and spontaneously hypertensive (SHR) decerebrate rats. Activation of mechanically sensitive fibers by passively stretching hindlimb muscle induced significantly greater increases in MAP and HR in SHR than WKY over a wide range of stimulus intensities. Activation of chemically sensitive fibers by administering capsaicin (0.01-1.00 microg/100 microl) into the hindlimb arterial supply induced increases in MAP that were significantly greater in SHR compared with WKY. However, HR responses to capsaicin were not different between the two groups at any dose. This data is consistent with the concept that the abnormal EPR control of MAP described previously in hypertension is mediated by both mechanoreflex and metaboreflex overactivity. In contrast, the previously reported alterations in the EPR control of HR in hypertension may be principally due to overactivity of the mechanically sensitive component of the reflex.

The glycine site-specific NMDA antagonist (+)-HA966 enhances the effect of morphine and reverses morphine tolerance via a spinal mechanism

Using the C-fibre reflex as a nociceptive response elicited by a wide range of stimulus intensities in the rat, we recently reported that a single treatment with (+)-HA966, a glycine site-specific NMDA receptor antagonist: (1) potentiates morphine antinociception; and (2) reverses an established morphine tolerance. We presently aimed at determining whether our observation was likely to result from a direct effect on the spinal cord or an indirect effect of supraspinal origin. In a 2 × 2 × 2 experimental design, we compared the effects of 5 mg/kg morphine in: (1) sham-operated rats or animals whose brainstems had been transected at the level of the obex; (2) rats that were implanted with pellets, either 150 mg morphine or placebo; and (3) animals injected either with saline or 10 mg/kg (+)-HA966. The control C-fibre reflexes were similar in all groups of animals. As compared to “non-tolerant” rats, the depressive effect of morphine was weaker in “morphine-tolerant” animals where the threshold did not change following morphine but the gain of the stimulus-response curve decreased, albeit to a significantly lesser extent than in the “non-tolerant” group. Whether in “non-tolerant” or “tolerant” groups, the effects of morphine were stronger in “obex-transected” than in “sham-operated” animals. In all groups, the effects of morphine were potentiated by the preliminary administration of (+)-HA966. However, in the “morphine-tolerant” group, the preliminary administration of (+)-HA966 was more potent in the “sham-operated” than in the “obex-transected” groups. Since overall effects were very similar in “sham-operated” and “obex-transected” animals, we concluded for our model that the critical site for the expression of the neuronal plastic changes associated with morphine tolerance lies in the spinal cord.

Task‐related changes in propriospinal excitation from hand muscles to human flexor carpi radialis motoneurones

This study addresses whether there is excitation from human hand muscles to flexor carpi radialis (FCR) motoneurones mediated through propriospinal circuits and, if so, whether it is used in specific motor tasks. Electrical stimuli to the ulnar nerve at wrist level produced an excitation in FCR motoneurones with characteristics typical of a propriospinally mediated effect: low threshold (0.6 x motor threshold (MT)), a group I effect that was not reproduced by purely cutaneous stimuli, long central delay (4.1 +/- 0.4 ms in single units), suppression when the stimulus intensity was increased, and facilitation of the corticospinal excitation at the premotoneuronal level. Ulnar-induced propriospinally mediated excitation was compared during selective voluntary contractions of the FCR and, at equivalent level of FCR EMG, during tasks in which the FCR was activated automatically in postural contractions rather than voluntarily (grip, pinching and pointing). The excitation was significantly greater during grip (and pinching) than during voluntary FCR contractions and pointing, whether measured in single motor units or tonic EMG activity, or whether the response to motor cortex stimulation was assessed as the compound motor-evoked potential or the corticospinal peak in single units. The discrepancy between the tasks appeared with ulnar intensities above 0.8 x MT and was then present across a wide range of stimulus intensities. This suggests a reduction in the corticospinal control of 'feedback inhibitory interneurones' mediating peripheral inhibition to propriospinal neurones during grip and pinching. The resulting more effective background excitation of propriospinal neurones by the peripheral input from hand muscles could contribute to stabilizing the wrist during grip.

Monday July 3, 2006 12:00-13:30 Hall 3a Platform Session Clinical Neurophysiology I

Abstract 1 V. Kimiskidis, 1 D. Kazis, 1 S. Papagiannopoulos, 1 G. Vasiliadis, 1 F. Zara, 2 X. Fitsioris, 2 G. Georgiadis, and 1 A. Kazis ( 1 III Neurological Department, Aristotle University of Thessaloniki, Greece , 2 Neurological Department, “Papageorgiou” Hospital, Thessaloniki, Greece ) Purpose: Transcranial magnetic stimulation (TMS) studies reflecting cortical and spinal inhibitory mechanisms. Previous studies of SP in epilepsy have generally provided divergent results. The objective of the present study is to investigate SP in untreated patients with idiopathic generalised epilepsy (IGE) using a novel methodological approach. Method: Fourteen patients with IGE (9 females, median age 19 yrs, range: 16–22) entered the study. Seven patients suffered from juvenile myoclonic epilepsy (JME), 4 patients from idiopathic epilepsy with generalised tonic–clonic seizures on awakening and the rest from other idiopathic generalised epilepsy syndromes not better defined. All electrophysiological examinations were performed at least 48 hours after the occurrence of an epileptic seizure so as to account for the confounding factor of postictal changes. Results were compared with those of a control group comprising 13 healthy, age-matched subjects. SPs were investigated as recently described [Kimiskidis et al. Exp Brain Res 2005;163:21–31]. First, SPs were elicited using a wide range of stimulus intensities (SIs) (from 5 to 100% maximum SI at 5% increments). At each SI, 4 SPs were obtained and the average value of SP duration was used to construct a stimulus/response (S/R) curve of SI vs SP. The resulting S/R curves were then fitted to a Boltzman function, the best-fit values of which were statistically compared between the patient and the control group. Results: The SP S/R curve of the patients was significantly different compared to the controls (p > 0.0001, F-test and AIC). In particular, the Max value of the patient's curve was 257.5 ± 3.98 ms vs 221.4 ± 2.89 ms in the controls (p < 0.0001) and V50 was 46.95 ± 0.71 vs 51.03 ± 0.57 (p < 0.0001) whereas slope was not significantly different (9.94 ± 0.58 vs 10.09 ± 0.46, p > 0.05). Conclusion: SP, a GABAB receptor mediated event, is prolonged in IGE. Basic neurophysiology experiments prove that GABAB IPSPs are more efficiently activated in the presence of GABAA receptor antagonists. Therefore, it could be hypothesised that a hypofunction of GABAA receptors in patients with IGE results in increased GABAB IPSPs reflected in prolonged SPs.

Tolerance to morphine analgesia: Evidence for stimulus intensity as a key factor and complete reversal by a glycine site-specific NMDA antagonist

N-methyl- d-aspartate (NMDA) receptors are widely involved in opioid tolerance. However, it is less clear whether NMDA receptor antagonists reverse already-established tolerance and whether the intensity of the nociceptive stimulus influences morphine tolerance. Three days after implantation of morphine or control pellets the effects of i.v. morphine and pre-administration of saline or (+)-HA966 (a glycine site-specific NMDA receptor antagonist), were studied on the C-fibre reflex elicited by a wide range of stimulus intensities. Morphine both increased the threshold and decreased the slope of the recruitment curve in the “non-tolerant” group of animals. In the “morphine-tolerant” group, the threshold did not change but the gain of the stimulus-response curve decreased. The expression of tolerance to morphine depended on the intensity of the stimulus, being maximal when threshold stimulus intensities were used but considerably less with supra-threshold stimulation. As expected, a single treatment with (+)-HA966, potentiated morphine antinociception in “non-tolerant” rats. However, in “morphine-tolerant” rats (+)-HA966 reversed established morphine tolerance and increased the antinociceptive effects of morphine. These results suggest that (+)-HA966 interfered with expression of morphine tolerance, and offered an encouraging therapeutic approach for pain management in opioid abusers.

NAD(P)H Fluorescence Transients after Synaptic Activity in Brain Slices: Predominant Role of Mitochondrial Function

Excitatory stimulation in hippocampal slices results in biphasic NAD(P)H fluorescence transients. Previous studies using differing stimulus protocols agreed that the oxidation phase is a consequence of mitochondrial metabolism, but the reduction phase has been attributed to (1) mitochondrial nicotinamide adenine dinucleotide (NADH) generation or (2) astrocytic glycolysis triggered by glutamate uptake. In an attempt to reconcile these two views, the present study examined NAD(P)H signals evoked by a wide range of stimulus durations (40 ms to 20 secs). A combination of ionotropic glutamate receptor (iGluR) antagonists (6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), 2-amino-5-phosphonopentanoic acid (APV)) virtually abolished responses to brief stimuli (40 to 200 ms, 50 Hz), but a significant fraction of the signal elicited by extended stimulation (20 secs, 32 Hz) was resistant to CNQX/APV. Glycolysis was inhibited by removal of glucose and addition of 2-deoxyglucose (2DG) (10 mmol/L) or iodoacetic acid (IAA, 1 mmol/L). Pyruvate was provided as an alternative substrate for oxidative phosphorylation and the A1 receptor antagonist 1,3-Dipropyl-8-cyclopentylxanthine (DPCPX) included to prevent decreases in synaptic efficacy. If sufficient pyruvate was supplied, responses to brief and extended stimuli were unaffected by glycolytic inhibition and not significantly reduced by an inhibitor of glucose uptake (3-O-methyl glucose, 3 mmol/L). When timed to arrive at the peak of overshoots generated by extended synaptic stimulation, brief pyruvate applications (10 mmol/L, 2 mins) had little effect on evoked NAD(P)H increases. Flavoprotein autofluorescence transients after extended stimuli matched (with inverted sign) NAD(P)H responses. Responses to extended stimuli were not reduced by a nonselective inhibitor of glutamate uptake DL-Threo-beta-benzyloxyaspartic acid (TBOA). These results suggest that NAD(P)H transients report mitochondrial dynamics, rather than recruitment of glycolytic metabolism, over a wide range of stimulus intensities.

Lorazepam-induced effects on silent period and corticomotor excitability

TMS studies on the CNS effects of benzodiazepines have provided contradictory results. The objective of this study is to describe the effects of lorazepam on silent period (SP) and corticomotor excitability. Twelve healthy male subjects (median age 35 years) were studied at baseline, following i.v. lorazepam administration and after reversal of the benzodiazepine effects with i.v. flumazenil. Lorazepam was given at a low-dose in one subject (0.0225 mg/kg bolus + 2 microg/kg/h infusion) and at a high-dose (0.045 mg/kg bolus + 2.6 microg/kg/h infusion) in the rest. Threshold (Thr) was measured at 1% steps. SPs were investigated with two complementary methods. First, SPs were elicited using a wide range of stimulus intensities (SIs) (from 5 to 100% maximum SI at 5% increments). At each SI, four SPs were obtained and the average value of SP duration was used to construct a stimulus/response (S/R) curve of SI versus SP .The resulting S/R curves were then fitted to a Boltzman function, the best-fit values of which were statistically compared for each experimental condition (i.e., baseline vs. lorazepam vs. flumazenil). Second, a large number of SPs (n=100) was elicited during each of the three experimental conditions using blocks of four stimuli with an intensity alternating between MT and 200% MT. This method was employed so as to reveal the dynamic, time-varying effects of lorazepam and flumazenil on SP duration at two stimulus intensity (SI) levels. MEP recruitment curves were constructed at rest and during activation and fitted to a Boltzman function the best-fit values of which were statistically compared for each experimental condition. Lorazepam at a low dose did not affect Thr, SP, or the active MEP recruitment curves. The high dose also had no effect on Thr and the active MEPs whereas the resting MEP recruitment curves were depressed post-lorazepam at the higher range of stimulus intensities. With regard to SP, the Max value of the S/R curve decreased from 251+/-4.6 ms at baseline to 215.2+/-3.1 ms post-lorazepam (P<0.01). V50 also decreased significantly (from 47.92+/-0.9% to 43.73+/-0.81%, P<0.01) whereas there was no significant change regarding slope and SP Thr. The statistical analysis of the SP S/R curves as well as the study of SPs at two SI levels revealed that lorazepam reduced SP duration when high intensity stimuli were used (>60%). In contrast, at low SIs a small increase in SP duration was noted post-drug. Enhancement of GABAergic inhibition by lorazepam results in a reduction of SP duration when high SIs is used. At the lower range of SIs, a small but statistically significant increase in SP duration is observed. The kinetic behavior of this phenomenon as well as the possible underlying mechanisms are discussed.