Nerve Injury-Induced Neuropathic Pain Causes Disinhibition of the Anterior Cingulate Cortex

Neuropathic pain caused by peripheral nerve injury is a debilitating neurological condition of high clinical relevance. On the cellular level, the elevated pain sensitivity is induced by plasticity of neuronal function along the pain pathway. Changes in cortical areas involved in pain processing contribute to the development of neuropathic pain. Yet, it remains elusive which plasticity mechanisms occur in cortical circuits. We investigated the properties of neural networks in the anterior cingulate cortex (ACC), a brain region mediating affective responses to noxious stimuli. We performed multiple whole-cell recordings from neurons in layer 5 (L5) of the ACC of adult mice after chronic constriction injury of the sciatic nerve of the left hindpaw and observed a striking loss of connections between excitatory and inhibitory neurons in both directions. In contrast, no significant changes in synaptic efficacy in the remaining connected pairs were found. These changes were reflected on the network level by a decrease in the mEPSC and mIPSC frequency. Additionally, nerve injury resulted in a potentiation of the intrinsic excitability of pyramidal neurons, whereas the cellular properties of interneurons were unchanged. Our set of experimental parameters allowed constructing a neuronal network model of L5 in the ACC, revealing that the modification of inhibitory connectivity had the most profound effect on increased network activity. Thus, our combined experimental and modeling approach suggests that cortical disinhibition is a fundamental pathological modification associated with peripheral nerve damage. These changes at the cortical network level might therefore contribute to the neuropathic pain condition.

Validation of stress magnetic resonance imaging of the canine stifle joint with and without an intact cranial cruciate ligament.

OBJECTIVE To validate use of stress MRI for evaluation of stifle joints of dogs with an intact or deficient cranial cruciate ligament (CrCL). SAMPLE 10 cadaveric stifle joints from 10 dogs. PROCEDURES A custom-made limb-holding device and a pulley system linked to a paw plate were used to apply axial compression across the stifle joint and induce cranial tibial translation with the joint in various degrees of flexion. By use of sagittal proton density-weighted MRI, CrCL-intact and deficient stifle joints were evaluated under conditions of loading stress simulating the tibial compression test or the cranial drawer test. Medial and lateral femorotibial subluxation following CrCL transection measured under a simulated tibial compression test and a cranial drawer test were compared. RESULTS By use of tibial compression test MRI, the mean ± SD cranial tibial translations in the medial and lateral compartments were 9.6 ± 3.7 mm and 10 ± 4.1 mm, respectively. By use of cranial drawer test MRI, the mean ± SD cranial tibial translations in the medial and lateral compartments were 8.3 ± 3.3 mm and 9.5 ± 3.5 mm, respectively. No significant difference in femorotibial subluxation was found between stress MRI techniques. Femorotibial subluxation elicited by use of the cranial drawer test was greater in the lateral than in the medial compartment. CONCLUSIONS AND CLINICAL RELEVANCE Both stress techniques induced stifle joint subluxation following CrCL transection that was measurable by use of MRI, suggesting that both methods may be further evaluated for clinical use.

Oviposition by a moth suppresses constitutive and herbivore-induced plant volatiles in maize

Plant volatiles function as important signals for herbivores, parasitoids, predators, and neighboring plants. Herbivore attack can dramatically increase plant volatile emissions in many species. However, plants do not only react to herbivore-inflicted damage, but also already start adjusting their metabolism upon egg deposition by insects. Several studies have found evidence that egg deposition itself can induce the release of volatiles, but little is known about the effects of oviposition on the volatiles released in response to subsequent herbivory. To study this we measured the effect of oviposition by Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) moths on constitutive and herbivore-induced volatiles in maize (Zea mays L.). Results demonstrate that egg deposition reduces the constitutive emission of volatiles and suppresses the typical burst of inducible volatiles following mechanical damage and application of caterpillar regurgitant, a treatment that mimics herbivory. We discuss the possible mechanisms responsible for reducing the plant's signaling capacity triggered by S. frugiperda oviposition and how suppression of volatile organic compounds can influence the interaction between the plant, the herbivore, and other organisms in its environment. Future studies should consider oviposition as a potential modulator of plant responses to insect herbivores. © 2011 Springer-Verlag.

Methylnitrosourea (MNU)-induced retinal degeneration and regeneration in the zebrafish: histological and functional characteristics.

Retinal degenerative diseases, e.g. retinitis pigmentosa, with resulting photoreceptor damage account for the majority of vision loss in the industrial world. Animal models are of pivotal importance to study such diseases. In this regard the photoreceptor-specific toxin N-methyl-N-nitrosourea (MNU) has been widely used in rodents to pharmacologically induce retinal degeneration. Previously, we have established a MNU-induced retinal degeneration model in the zebrafish, another popular model system in visual research. A fascinating difference to mammals is the persistent neurogenesis in the adult zebrafish retina and its regeneration after damage. To quantify this observation we have employed visual acuity measurements in the adult zebrafish. Thereby, the optokinetic reflex was used to follow functional changes in non-anesthetized fish. This was supplemented with histology as well as immunohistochemical staining for apoptosis (TUNEL) and proliferation (PCNA) to correlate the developing morphological changes. In summary, apoptosis of photoreceptors occurs three days after MNU treatment, which is followed by a marked reduction of cells in the outer nuclear layer (ONL). Thereafter, proliferation of cells in the inner nuclear layer (INL) and ONL is observed. Herein, we reveal that not only a complete histological but also a functional regeneration occurs over a time course of 30 days. Now we illustrate the methods to quantify and follow up zebrafish retinal de- and regeneration using MNU in a video-format.

Characterization of the role of the RNA-binding protein FUS in the DNA damage response

FUS/TLS (fused in sarcoma/translocated in liposarcoma), a ubiquitously expressed RNA-binding protein, has been linked to a variety of cellular processes, including RNA metabolism, microRNA biogenesis and DNA repair. However, the precise cellular function of FUS remains unclear. Recently, mutations in the FUS gene have been found in ∼5% of familial Amyotrophic Lateral Sclerosis, a neurodegenerative disorder characterized by the dysfunction and death of motor neurons. Since MEFs and B-lymphocytes derived from FUS knockdown mice display major sensitivity to ionizing radiation and chromosomal aberrations [1,2], we are investigating the effects of DNA damage both in the presence or in the absence of FUS. To this purpose, we have generated a SH-SY5Y human neuroblastoma cell line expressing a doxycycline-induced shRNA targeting FUS, which specifically depletes the protein. We have found that FUS depletion induces an activation of the DNA damage response (DDR). However, treatment with genotoxic agents did not induce any strong changes in ATM (Ataxia Telangiectasia Mutated)-mediated DDR signaling. Interestingly, genotoxic treatment results in changes in the subcellular localization of FUS in normal cells. We are currently exploring on one hand the mechanism by which FUS depletion leads to DNA damage, and on the other the functional significance of FUS relocalization after genotoxic stress.

Refinement of tourniquet-induced peripheral ischemia/reperfusion injury in rats: comparison of 2 h vs 24 h reperfusion.

Prolonged ischemia of skeletal muscle tissue, followed by reperfusion, leads to ischemia/reperfusion injury (IRI), which is a feared local and systemic inflammatory reaction. With respect to the 3Rs, we wanted to determine which parameters for assessment of IRI require a reperfusion time of 24 h and for which 2 h of reperfusion are sufficient. Rats were subjected to 3 h of hind limb ischemia and 2 h or 24 h of reperfusion. Human plasma derived C1 inhibitor was used as a drug to prevent reperfusion injury. For 2 h of reperfusion the rats stayed under anesthesia throughout (severity grade 1), whereas for 24 h they were awake under analgesia during reperfusion (grade 2). The femoral artery was clamped and a tourniquet was placed, under maintenance of venous return. C1 esterase inhibitor was systemically administered 5 min before the induction of ischemia. No differences in local muscle edema formation and depositions of immunoglobulin G and immunoglobulin M were observed between 2 h and 24 h (P > 0.05), whereas lung edema was only observed after 24 h. Muscle viability was significantly lower after 24 h vs 2 h reperfusion (P < 0.05). Increased plasma creatine kinase (CK)-MM and platelet-derived growth factor (PDGF)-bb could be detected after 2 h, but not after 24 h of reperfusion. By contrast, depositions of C3b/c and fibrin in muscle were only detected after 24 h (P < 0.001). In conclusion, for a first screening of drug candidates to reduce IRI, 2 h reperfusions are sufficient, and these reduce the severity of the animal experiment. Twenty-four-hour reperfusions are only needed for in-depth analysis of the mechanisms of IRI, including lung damage.

Rhythmic synaptic activity induced by mechanical injury of rat CA3 hippocampal area.

Mechanical injury of the CNS frequently results from accidents but also occurs in the course of neurosurgical interventions. A great variety of anatomical and physiological changes have been described to evolve after a brain trauma yet only little is known about processes that occur during a trauma. In the present study, I obtained whole-cell patch clamp recordings from pyramidal cells in hippocampal slice cultures while mechanically lesioning the CA3 area. Electrophysiological analysis revealed that traumatic injury massively increased excitatory and inhibitory synaptic activity in the entire CA3 region. Cutting the CA3 region induced highly rhythmic excitatory postsynaptic currents (EPSCs) that reached frequencies of around 70 Hz. Blocking voltage-dependent sodium channels with tetrodotoxin prevented the increase in synaptic activity and injury-induced neurotransmitter release in CA3 remote from the lesion site. With fast synaptic transmission blocked only neurons in the immediate vicinity of a lesion depolarized and fired action potentials upon mechanical damage. I hence suggest that mechanical injury damages the membrane and induces action potential firing in only a small population of neurons. This activity is then propagated throughout the undamaged CA3 network inducing highly rhythmic discharges. Thus mechanical brain injury initiates immediate functional changes that exceed the lesion site.

Induced Immunity Against Belowground Insect Herbivores- Activation of Defenses in the Absence of a Jasmonate Burst

Roots respond dynamically to belowground herbivore attack. Yet, little is known about the mechanisms and ecological consequences of these responses. Do roots behave the same way as leaves, or do the paradigms derived from aboveground research need to be rewritten? This is the central question that we tackle in this article. To this end, we review the current literature on induced root defenses and present a number of experiments on the interaction between the root herbivore Diabrotica virgifera and its natural host, maize. Currently, the literature provides no clear evidence that plants can recognize root herbivores specifically. In maize, mild mechanical damage is sufficient to trigger a root volatile response comparable to D. virgifera induction. Interestingly, the jasmonate (JA) burst, a highly conserved signaling event following leaf attack, is consistently attenuated in the roots across plant species, from wild tobacco to Arabidopsis. In accordance, we found only a weak JA response in D. virgifera attacked maize roots. Despite this reduction in JA-signaling, roots of many plants start producing a distinct suite of secondary metabolites upon attack and reconfigure their primary metabolism. We, therefore, postulate the existence of additional, unknown signals that govern induced root responses in the absence of a jasmonate burst. Surprisingly, despite the high phenotypic plasticity of plant roots, evidence for herbivore-induced resistance below ground is virtually absent from the literature. We propose that other defensive mechanisms, including resource reallocation and compensatory growth, may be more important to improve plant immunity below ground.

Two enzymes responsible for the formation of herbivore-induced volatiles of maize, the methyltransferase AAMT1 and the terpene synthase TPS23, are regulated by a similar signal transduction pathway

Herbivore-induced volatiles play an important role in the indirect defense of plants. After herbivore damage, volatiles are released from the plant and can attract herbivore enemies that protect the plant from additional damage. The herbivore-induced volatile blend is complex and usually consists of mono- and sesquiterpenes, aromatic compounds, and indole. Although these classes of compounds are generally produced at different times after herbivore damage, the release of the terpene (E)-β-caryophyllene and the aromatic ester methyl anthranilate appear to be tightly coordinated. We have studied the herbivore induction patterns of two terpene synthases from Zea mays L. (Poaceae), TPS23 and TPS10, as well as S-adenosyl-L-methionine:anthranilic acid carboxyl methyltransferases (AAMT1), which are critical for the production of terpenes and anthranilate compounds, respectively. The transcript levels of tps23 and aamt1 displayed the same kinetics after damage by the larvae of Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae), and showed the same organ-specific and haplotype-specific expression patterns. Despite its close functional relation to TPS23, the terpene synthase TPS10 is not expressed in roots and does not display the haplotype-specific expression pattern. The results indicate that the same JA-mediated signaling cascade maycontrol the production of both the terpene (E)-β-caryophyllene and aromatic ester methyl anthranilate.

Temporal variability of damage potential on roads as a conceptual contribution towards a short-term avalanche risk simulation

The fatality risk caused by avalanches on road networks can be analysed using a long-term approach, resulting in a mean value of risk, and with emphasis on short-term fluctuations due to the temporal variability of both, the hazard potential and the damage potential. In this study, the approach for analysing the long-term fatality risk has been adapted by modelling the highly variable short-term risk. The emphasis was on the temporal variability of the damage potential and the related risk peaks. For defined hazard scenarios resulting from classified amounts of snow accumulation, the fatality risk was calculated by modelling the hazard potential and observing the traffic volume. The avalanche occurrence probability was calculated using a statistical relationship between new snow height and observed avalanche releases. The number of persons at risk was determined from the recorded traffic density. The method resulted in a value for the fatality risk within the observed time frame for the studied road segment. The long-term fatality risk due to snow avalanches as well as the short-term fatality risk was compared to the average fatality risk due to traffic accidents. The application of the method had shown that the long-term avalanche risk is lower than the fatality risk due to traffic accidents. The analyses of short-term avalanche-induced fatality risk provided risk peaks that were 50 times higher than the statistical accident risk. Apart from situations with high hazard level and high traffic density, risk peaks result from both, a high hazard level combined with a low traffic density and a high traffic density combined with a low hazard level. This provided evidence for the importance of the temporal variability of the damage potential for risk simulations on road networks. The assumed dependence of the risk calculation on the sum of precipitation within three days is a simplified model. Thus, further research is needed for an improved determination of the diurnal avalanche probability. Nevertheless, the presented approach may contribute as a conceptual step towards a risk-based decision-making in risk management.

An improved simple rat model for global cerebral ischaemia by induced cardiac arrest

OBJECTIVES Cerebral hypoxic-ischaemic injury following cardiac arrest is a devastating disease affecting thousands of patients each year. There is a complex interaction between post-resuscitation injury after whole-body ischaemia-reperfusion and cerebral damage which cannot be explored in in vitro systems only; there is a need for animal models. In this study, we describe and evaluate the feasibility and efficiency of our simple rodent cardiac arrest model. METHODS Ten wistar rats were subjected to 9 and 10 minutes of cardiac arrest. Cardiac arrest was introduced with a mixture of the short-acting beta-blocking drug esmolol and potassium chloride. RESULTS All animals could be resuscitated within 1 minute, and survived until day 5.General health score and neurobehavioural testing indicated substantial impairment after cardiac arrest, without differences between groups. Histological examination of the hippocampus CA1 segment, the most vulnerable segment of the cerebrum, demonstrated extensive damage in the cresyl violet staining, as well as in the Fluoro-Jade B staining and in the Iba-1 staining, indicating recruitment of microglia after the hypoxic-ischaemic event. Again, there were no differences between the 9- and 10-minute cardiac arrest groups. DISCUSSION We were able to establish a simple and reproducible 9- and 10-minute rodent cardiac arrest models with a well-defined no-flow-time. Extensive damage can be found in the hippocampus CA1 segment. The lack of difference between 9- and 10-minute cardiac arrest time in the neuropsychological, the open field test and the histological evaluations is mainly due to the small sample size.

Herbivore-Induced Plant Volatiles Can Serve as Host Location Cues for a Generalist and a Specialist Egg Parasitoid

Herbivore-induced plant volatiles are important host finding cues for larval parasitoids, and similarly, insect oviposition might elicit the release of plant volatiles functioning as host finding cues for egg parasitoids. We hypothesized that egg parasitoids also might utilize HIPVs of emerging larvae to locate plants with host eggs. We, therefore, assessed the olfactory response of two egg parasitoids, a generalist, Trichogramma pretiosum (Tricogrammatidae), and a specialist, Telenomus remus (Scelionidae) to HIPVs. We used a Y-tube olfactometer to tests the wasps’ responses to volatiles released by young maize plants that were treated with regurgitant from caterpillars of the moth Spodoptera frugiperda (Noctuidae) or were directly attacked by the caterpillars. The results show that the generalist egg parasitoid Tr. pretiosum is innately attracted by volatiles from freshly-damaged plants 0–1 and 2–3 h after regurgitant treatment. During this interval, the volatile blend consisted of green leaf volatiles (GLVs) and a blend of aromatic compounds, mono- and homoterpenes, respectively. Behavioral assays with synthetic GLVs confirmed their attractiveness to Tr. pretiosum. The generalist learned the more complex volatile blends released 6–7 h after induction, which consisted mainly of sesquiterpenes. The specialist T. remus on the other hand was attracted only to volatiles emitted from fresh and old damage after associating these volatiles with oviposition. Taken together, these results strengthen the emerging pattern that egg and larval parasitoids behave in a similar way in that generalists can respond innately to HIPVs, while specialists seems to rely more on associative learning.

Biochemical MRI predicts hip osteoarthritis in an experimental ovine femoroacetabular impingement model

BACKGROUND Cam-type femoroacetabular impingement (FAI) resulting from an abnormal nonspherical femoral head shape leads to chondrolabral damage and is considered a cause of early osteoarthritis. A previously developed experimental ovine FAI model induces a cam-type impingement that results in localized chondrolabral damage, replicating the patterns found in the human hip. Biochemical MRI modalities such as T2 and T2* may allow for evaluation of the cartilage biochemistry long before cartilage loss occurs and, for that reason, may be a worthwhile avenue of inquiry. QUESTIONS/PURPOSES We asked: (1) Does the histological grading of degenerated cartilage correlate with T2 or T2* values in this ovine FAI model? (2) How accurately can zones of degenerated cartilage be predicted with T2 or T2* MRI in this model? METHODS A cam-type FAI was induced in eight Swiss alpine sheep by performing a closing wedge intertrochanteric varus osteotomy. After ambulation of 10 to 14 weeks, the sheep were euthanized and a 3-T MRI of the hip was performed. T2 and T2* values were measured at six locations on the acetabulum and compared with the histological damage pattern using the Mankin score. This is an established histological scoring system to quantify cartilage degeneration. Both T2 and T2* values are determined by cartilage water content and its collagen fiber network. Of those, the T2* mapping is a more modern sequence with technical advantages (eg, shorter acquisition time). Correlation of the Mankin score and the T2 and T2* values, respectively, was evaluated using the Spearman's rank correlation coefficient. We used a hierarchical cluster analysis to calculate the positive and negative predictive values of T2 and T2* to predict advanced cartilage degeneration (Mankin ≥ 3). RESULTS We found a negative correlation between the Mankin score and both the T2 (p < 0.001, r = -0.79) and T2* values (p < 0.001, r = -0.90). For the T2 MRI technique, we found a positive predictive value of 100% (95% confidence interval [CI], 79%-100%) and a negative predictive value of 84% (95% CI, 67%-95%). For the T2* technique, we found a positive predictive value of 100% (95% CI, 79%-100%) and a negative predictive value of 94% (95% CI, 79%-99%). CONCLUSIONS T2 and T2* MRI modalities can reliably detect early cartilage degeneration in the experimental ovine FAI model. CLINICAL RELEVANCE T2 and T2* MRI modalities have the potential to allow for monitoring the natural course of osteoarthrosis noninvasively and to evaluate the results of surgical treatments targeted to joint preservation.

First Metatarsophalangeal Joint Degeneration: Arthroscopic Treatment

Arthroscopic treatment of hallux rigidus is appropriate after failed nonoperative treatment. Debridement with cheilectomy, and fusion are the main indications for arthroscopic treatment of hallux rigidus. If the cartilage damage is extensive and the patient has consented, then a fusion is performed at the same sitting.

T1ρ MRI detects cartilage damage in asymptomatic individuals with a cam deformity

Hips with a cam deformity are at risk for early cartilage degeneration, mainly in the anterolateral region of the joint. T1ρ MRI is a described technique for assessment of proteoglycan content in hyaline cartilage and subsequently early cartilage damage. In this study, 1.5 Tesla T1ρ MRI was performed on 20 asymptomatic hips with a cam deformity and compared to 16 healthy control hips. Cam deformity was defined as an alpha angle at 1:30 o'clock position over 60° and/or at 3:00 o'clock position over 50.5°. Hip cartilage was segmented and divided into four regions of interest (ROIs): anterolateral, anteromedial, posterolateral and posteromedial quadrants. Mean T1ρ value of the entire weight bearing cartilage in hips with a cam deformity (34.0 ± 4.6 ms) was significantly higher compared to control hips (31.3 ± 3.2 ms, p = 0.050). This difference reached significance in the anterolateral (p = 0.042) and posteromedial quadrants (p = 0.041). No significant correlation between the alpha angle and T1ρ values was detected. The results indicate cartilage damage occurs in hips with a cam deformity before symptoms occur. A significant difference in T1ρ values was found in the anterolateral quadrant, the area of direct engagement of the deformity, and in the posteromedial quadrant. To conclude, T1ρ MRI can detect early chondral damage in asymptomatic hips with a cam deformity. This article is protected by copyright. All rights reserved.