Progenitor Hyperpolarization Regulates the Sequential Generation of Neuronal Subtypes in the Developing Neocortex.

During corticogenesis, ventricular zone progenitors sequentially generate distinct subtypes of neurons, accounting for the diversity of neocortical cells and the circuits they form. While activity-dependent processes are critical for the differentiation and circuit assembly of postmitotic neurons, how bioelectrical processes affect nonexcitable cells, such as progenitors, remains largely unknown. Here, we reveal that, in the developing mouse neocortex, ventricular zone progenitors become more hyperpolarized as they generate successive subtypes of neurons. Experimental in vivo hyperpolarization shifted the transcriptional programs and division modes of these progenitors to a later developmental status, with precocious generation of intermediate progenitors and a forward shift in the laminar, molecular, morphological, and circuit features of their neuronal progeny. These effects occurred through inhibition of the Wnt-beta-catenin signaling pathway by hyperpolarization. Thus, during corticogenesis, bioelectric membrane properties are permissive for specific molecular pathways to coordinate the temporal progression of progenitor developmental programs and thus neocortical neuron diversity.

Endogenous erythropoietin signaling regulates migration and laminar positioning of upper-layer neurons in the developing neocortex.

Erythropoietin (EPO), the hypoxia-inducible hematopoietic hormone, has well-established neuroprotective/neurotrophic roles in the developing central nervous system and the therapeutic potential of EPO has been widely explored in clinical studies for the treatment of perinatal hypoxic brain lesion, as well as prematurity. Here, we reveal that both EPO and Epo receptor (EPOR) are expressed in the developing rat somatosensory cortex during radial migration and laminar positioning of granular and supragranular neurons. Experimental deregulation of EPO signaling using genetic approaches results in aberrant migration, as well as permanent neuronal misplacement leading to abnormal network activity and protracted sensory behavioral deficits. We identify ERK as the downstream effector of the EPO signaling pathway for neuronal migration. These findings reveal a crucial role for endogenous EPO signaling in neuronal migration, and offer important insights for understanding how the temporary deregulation of EPO could result in migration defects that lead to abnormal behavior in the adult.

Genetic Etiology of Nonsyndromic Hearing Loss in Hungarian Patients.

Hearing loss is the most prevalent sensory disorder worldwide. The majority of congenital nonsyndromic hearing loss (NSHL) cases are caused by hereditary factors. Previously, the majority of NSHL studies focused on the GJB2 gene; however, with the availability of next-generation sequencing (NGS) methods, the number of novel variants associated with NSHL has increased. The purpose of this study was to design effective genetic screening for a Hungarian population based on a pilot study with 139 NSHL patients. A stepwise, comprehensive genetic approach was developed, including bidirectional capillary sequencing, multiplex ligation-dependent probe amplification (MLPA), and an NGS panel of 108 hearing loss genes. With our results, a genetic diagnosis was possible for 92 patients. Sanger sequencing and MLPA identified the genetic background of 50% of these diagnosed cases, and the NGS panel identified another 16%. The vast majority (92%) of the diagnosed cases showed autosomal recessive inheritance and 76% were attributed to GJB2. The implementation of this stepwise analysis markedly increased our diagnostic yield and proved to be cost-effective as well.

A new method of preoperative assessment of correct electrode array alignment based on post-operative measurements in a cochlear implanted cohort.

PURPOSE: During cochlear implantation surgery, a range of complications may occur such as tip fold-over. We recently developed a method to estimate the insertion orientation of the electrode array. The aim of the study was to determine the optimal angle of orientation in a cohort of cochlear implanted patients. METHODS: On eighty-five CT scans (80 uncomplicated insertions and 5 cases with tip fold-over), location of the electrode array's Insertion Guide (IG), Orientation marker (OM) and two easily identifiable landmarks (the round window (RW) and the incus short process (ISP)) were manually marked. The angle enclosed by ISP-RW line and the Cochlear™ Slim Modiolar electrode array's OM line determined the electrode array insertion angle. RESULTS: The average insertion angle was 45.0-47.2° ± 10.4-12° SD and was validated with 98% confidence interval. Based on the measurements obtained, patients' sex and age had no impact on the size of this angle. Although the angles of the tip fold-over cases (44.9°, 46.9°, 34.2°, 54.3°, 55.9°) fell within this average range, the further it diverted from the average it increased the likelihood for tip fold-over. CONCLUSION: Electrode array insertion in the individually calculated angle relative to the visible incus short process provides a useful guide for the surgeon when aiming for the optimal angle, and potentially enhances good surgical outcomes. Our results show that factors other than the orientation angle may additionally contribute to failures in implantation when the Slim Modiolar electrode is used.

Transplanted Embryonic Neurons Improve Functional Recovery by Increasing Activity in Injured Cortical Circuits.

Transplantation of appropriate neuronal precursors after injury is a promising strategy to reconstruct cortical circuits, but the efficiency of these approaches remains limited. Here, we applied targeted apoptosis to selectively ablate layer II/III pyramidal neurons in the rat juvenile cerebral cortex and attempted to replace lost neurons with their appropriate embryonic precursors by transplantation. We demonstrate that grafted precursors do not migrate to replace lost neurons but form vascularized clusters establishing reciprocal synaptic contacts with host networks and show functional integration. These heterotopic neuronal clusters significantly enhance the activity of the host circuits without causing epileptic seizures and attenuate the apoptotic injury-induced functional deficits in electrophysiological and behavioral tests. Chemogenetic activation of grafted neurons further improved functional recovery, and the persistence of the graft was necessary for maintaining restored functions in adult animals. Thus, implanting neuronal precursors capable to form synaptically integrated neuronal clusters combined with activation-based approaches represents a useful strategy for helping long-term functional recovery following brain injury.

Late simultaneous carcinomatous meningitis, temporal bone infiltrating macro-metastasis and disseminated multi-organ micro-metastases presenting with mono-symptomatic vertigo - a clinico-pathological case reporT.

BACKGROUND AND PURPOSE: Although vertigo is one of the most common complaints, intracranial malignant tumors rarely cause sudden asymmetry between the tone of the vestibular peripheries masquerading as a peripheral-like disorder. Here we report a case of simultaneous temporal bone infiltrating macro-metastasis and disseminated multi-organ micro-metastases presenting as acute unilateral vestibular syndrome, due to the reawakening of a primary gastric signet ring cell carcinoma. Purpose - Our objective was to identify those pathophysiological steps that may explain the complex process of tumor reawakening, dissemination. The possible causes of vestibular asymmetry were also traced. METHODS: A 56-year-old male patient's interdisciplinary medical data had been retrospectively analyzed. Original clinical and pathological results have been collected and thoroughly reevaluated, then new histological staining and immunohistochemistry methods have been added to the diagnostic pool. RESULTS: During the autopsy the cerebrum and cerebellum was edematous. The apex of the left petrous bone was infiltrated and destructed by a tumor mass of 2x2 cm in size. Histological reexamination of the original gastric resection specimen slides revealed focal submucosal tumorous infiltration with a vascular invasion. By immunohistochemistry mainly single infiltrating tumor cells were observed with Cytokeratin 7 and Vimentin positivity and partial loss of E-cadherin staining. The subsequent histological examination of necropsy tissue specimens confirmed the disseminated, multi-organ microscopic tumorous invasion. Discussion - It has been recently reported that the expression of Vimentin and the loss of E-cadherin is significantly associated with advanced stage, lymph node metastasis, vascular and neural invasion and undifferentiated type with p<0.05 significance. As our patient was middle aged and had no immune-deficiency, the promoting factor of the reawakening of the primary GC malignant disease after a 9-year-long period of dormancy remained undiscovered. The organ-specific tropism explained by the "seed and soil" theory was unexpected, due to rare occurrence of gastric cancer to metastasize in the meninges given that only a minority of these cells would be capable of crossing the blood brain barrier. CONCLUSION: Patients with past malignancies and new onset of neurological symptoms should alert the physician to central nervous system involvement, and the appropriate, targeted diagnostic and therapeutic work-up should be established immediately. Targeted staining with specific antibodies is recommended. Recent studies on cell lines indicate that metformin strongly inhibits epithelial-mesenchymal transition of gastric cancer cells. Therefore, further studies need to be performed on cases positive for epithelial-mesenchymal transition.

Review of electrode placement with the Slim Modiolar Electrode: identification and management.

Background - Several cochlear implant recipients experience functionality loss due to electrode array mal-positioning. The application of delicate perimodiolar electrodes has many electrophysiological advantages, however, these profiles may be more susceptible to tip fold-over. Purpose - The prompt realization of such complication following electrode insertion would be auspicious, thus the electrode could be possibly repositioned during the same surgical procedure. Methods - The authors present three tip fold-over cases, experienced throughout their work with Slim Modiolar Electrode implants. Implantations were performed through the round window approach, by a skilled surgeon. Standard intraoperative measurements (electric integrity, neural response telemetry, and electrical stapedial reflex threshold tests) were successfully completed. The electrode position was controlled by conventional radiography on the first postoperative day. Results - Tip fold-over was not tactilely sensated by the surgeon. Our subjects revealed normal intraoperative telemetry measurements, only the postoperative imaging showed the tip fold-over. Due to the emerging adverse perception of constant beeping noise, the device was replaced by a CI512 implant after 6 months in one case. In the two remaining cases, the electrode array was reloaded into a back-up sheath, and reinserted into the scala tympani successfully through an extended round window approach. Discussion - Future additional studies using the spread of excitation or electric field imaging may improve test reliability. As all of these measurements are still carried out following electrode insertion, real-time identification, unfortunately, remains questionable. Conclusion - Tip fold-over could be reliably identified by conventional X-ray imaging. By contrast, intraoperative electrophysiology was not sufficiently sensitive to reveal it.

Electrophysiological Evidence for the Development of a Self-Sustained Large-Scale Epileptic Network in the Kainate Mouse Model of Temporal Lobe Epilepsy.

Most research on focal epilepsy focuses on mechanisms of seizure generation in the primary epileptic focus (EF). However, neurological deficits that are not directly linked to seizure activity and that may persist after focus removal are frequent. The recruitment of remote brain regions of an epileptic network (EN) is recognized as a possible cause, but a profound lack of experimental evidence exists concerning their recruitment and the type of pathological activities they exhibit. We studied the development of epileptic activities at the large-scale in male mice of the kainate model of unilateral temporal lobe epilepsy using high-density surface EEG and multiple-site intracortical recordings. We show that, along with focal spikes and fast ripples that remain localized to the injected hippocampus (i.e., the EF), a subpopulation of spikes that propagate across the brain progressively emerges even before the expression of seizures. The spatiotemporal propagation of these generalized spikes (GSs) is highly stable within and across animals, defining a large-scale EN comprising both hippocampal regions and frontal cortices. Interestingly, GSs are often concomitant with muscular twitches. In addition, while fast ripples are, as expected, highly frequent in the EF, they also emerge in remote cortical regions and in particular in frontal regions where GSs propagate. Finally, we demonstrate that these remote interictal activities are dependent on the focus in the early phase of the disease but continue to be expressed after focus silencing at later stages. Our results provide evidence that neuronal networks outside the initial focus are progressively altered during epileptogenesis.SIGNIFICANCE STATEMENT It has long been held that the epileptic focus is responsible for triggering seizures and driving interictal activities. However, focal epilepsies are associated with heterogeneous symptoms, calling into question the concept of a strictly focal disease. Using the mouse model of hippocampal sclerosis, this work demonstrates that focal epilepsy leads to the development of pathological activities specific to the epileptic condition, notably fast ripples, that appear outside of the primary epileptic focus. Whereas these activities are dependent on the focus early in the disease, focus silencing fails to control them in the chronic stage. Thus, dynamical changes specific to the epileptic condition are built up outside of the epileptic focus along with disease progression, which provides supporting evidence for network alterations in focal epilepsy.

Multimodal MRI Imaging of Apoptosis-Triggered Microstructural Alterations in the Postnatal Cerebral Cortex.

Prematurely born children often develop neurodevelopmental delay that has been correlated with reduced growth and microstructural alterations in the cerebral cortex. Much research has focused on apoptotic neuronal cell death as a key neuropathological features following preterm brain injuries. How scattered apoptotic death of neurons may contribute to microstructural alterations remains unknown. The present study investigated in a rat model the effects of targeted neuronal apoptosis on cortical microstructure using in vivo MRI imaging combined with neuronal reconstruction and histological analysis. We describe that mild, targeted death of layer IV neurons in the developing rat cortex induces MRI-defined metabolic and microstructural alterations including increased cortical fractional anisotropy. Delayed architectural modifications in cortical gray matter and myelin abnormalities in the subcortical white matter such as hypomyelination and microglia activation follow the acute phase of neuronal death and axonal degeneration. These results establish the link between mild cortical apoptosis and MRI-defined microstructure changes that are reminiscent to those previously observed in preterm babies.

[Possibilities for residual hearing preservation with Nucleus CI532 Slim Modiolar electrode array. Case report]. / A maradványhallás megorzésének lehetoségei cochlearis implantáció során Nucleus CI532 Slim Modiolar elektródasorral.

During the rehabilitation of hearing-impaired patients, the preservation of residual acoustic hearing following cochlear implantation by minimizing the implantation trauma allows for improved hearing performance. To achieve this, minimally invasive, soft surgery methods and thinner, atraumatic electrodes were required. In our present study, we reported a case where Cochlear® Nucleus CI532 Slim Modiolar electrode was implanted in a patient with residual hearing. Our aim was to study the possible preservation of postoperative acoustic residual hearing by audiological monitoring. Since childhood, due to her congenital hearing loss, she has been wearing a conventional, airborne hearing correction device on both ears. Six months before cochlear implantation, we measured the progression on both sides of the hearing loss, so we decided to perform cochlear implantation. The patient had residual hearing on both ears prior to surgery thus the Cochlear® Nucleus CI532 Slim Modiolar Implant was used. The minimally invasive surgery was performed on the patient's right ear through the round window approach. Compared to the preoperative hearing threshold (average 85 dBHL) in the 4th postoperative week, an initial hearing threshold progression of 20-25 dBHL was observed between 0.25 and 1.0 kHz, while of 5-10 dBHL between 2.0-4.0 kHz. Hearing threshold measured in the 6th month showed a slight progression in the range above 1 kHz, but improved by the 12th month, to the results achieved at the 4th week. The effects of cochlear implantation on residual hearing have been studied in numerous studies, in which several key surgical and technical factors have been identified. Nucleus CI532 is a Slim Modiolar electrode profile that is close to the modiolus, so it is expected to have a lower endocochlear hydrodynamic load since it lies in the covering of the osseus spiral lamina, thus less influencing the dynamics of the basilar membrane. However, the perimodiolar location of the electrode array allows the adjacent nerve elements of the spiral ganglion to be stimulated with a lower electrical intensity and a reduced surface that may be neuroprotective. Preservation of acoustic residual hearing following cochlear implantation improves the patient's speech perception and the sound localization skills, particularly in difficult circumstances. Long-term residual hearing preservation may also be of great importance in the subsequent feasibility for regenerative procedures and drug treatments. Orv Hetil. 2018; 159(41): 1680-1688.

[The role of laryngeal electromyography in the diagnosis of vocal cord movement disorders]. / A laryngealis electromyographia szerepe a hangszalag-mozgászavarok diagnosztikájában és az alkalmazott kezelés kiválasztásában.

The development of the therapeutic possibilities of vocal cord immobility necessitated the parallel renewal of diagnostic methods. In the last years, laryngeal electromyography, which was first introduced more than 70 years ago, has been re-discovered. After reviewing the international literature and their own experience, the authors present the indications, technical requirements, method and, particularly, the evaluation of the results of this procedure. Laryngeal electromyography makes the differentiation between mechanical fixation and immobility with neurological origin of the vocal folds possible. In case of laryngeal paralysis/paresis it also evaluates objectively the severity of neural injury, the prognosis of the disease and the necessity of any glottis-widening procedure. The widespread application of dynamic rehabilitation interventions is not conceivable without the routine application of laryngeal electromyography, so this sensitive diagnostic tool has to be introduced in all laryngological centers. Orv Hetil. 2018; 159(8): 303-311.

The chemical inducer, BTH (benzothiadiazole) and root colonization by mycorrhizal fungi (Glomus spp.) trigger resistance against white rot (Sclerotinia sclerotiorum) in sunflower.

White rot caused by Sclerotinia sclerotiorum (SS) is one of the most devastating plant diseases of sunflower. Controlling this pathogen by available tools hardly result in acceptable control. The aim of this study was to elucidate the effects of plant resistance inducers, BTH (benzothiadiazole in Bion 50 WG) and arbuscular mycorrhizal fungi (AMF) on disease development of white rot in three sunflower genotypes. Defence responses were characterized by measuring the disease severity and identifying cellular/histological reactions (e.g. autofluorescence) of host plants upon infection. Depending on the host genotype, a single application of inducers reduced disease symptoms. Histological examination of host responses revealed that BTH and/or AMF pre-treatments significantly impeded the development of pathogenic hyphae in Iregi szürke csíkos and P63LE13 sunflower plants and it was associated with intensive autofluorescence of cells. Both localized and systemic induction of resistance was observed. Importantly, the frequency of mycorrhization of hybrid P63LE13 and PR64H41 was significantly increased upon BTH treatment, so it had a positive effect on the formation of plant-mycorrhiza interactions in sunflower. To our knowledge, this is the first report on the additive effect of BTH on mycorrhization and the positive effect of these inducers against SS in sunflower.

[Pediatric hearing rehabilitation with the Baha® Attract implant system]. / Fiatalkori hallásrehabilitáció Baha® Attract implantátumrendszerrel.

INTRODUCTION: Baha® Attract is a new transcutaneous bone-conduction hearing aid, which is more preferable in childhood than the conventional percutaneous systems. AIM: Our aim was to demonstrate the possibilities of application in childhood. METHOD: Eight children have undergone surgeries (mean age of 13.2 ± 3.2 years; "posterosuperior" incision technique, 5 mm implants). The thickness of the skull bone was determined in 72 children (1-8 years old) at the recommended implant site, based on CT scans. RESULTS: The average duration of surgeries was 30 minutes. There were no intra- and postoperative complications observed. Sound processors were fitted at the postoperative 4th week. Hearing measurements proved 51.58±11.22SD dBHL gain in warble tone thresholds, and 43.3 ± 16.02 SD dB in speech discrimination thresholds. The skull bone thickness was measured as 3.39 ± 1.05 SD mm. CONCLUSION: The Baha Attract system is a new tool for hearing rehabilitation in pediatric population. Preoperative CT provides valuable knowledge about skull bone thickness. Orv. Hetil., 2017, 158(8), 304-310.

Astrocytes spatially restrict VEGF signaling by polarized secretion and incorporation of VEGF into the actively assembling extracellular matrix.

The spatial organization of vascular endothelial growth factor (VEGF) signaling is a key determinant of vascular patterning during development and tissue repair. How VEGF signaling becomes spatially restricted and the role of VEGF secreting astrocytes in this process remains poorly understood. Using a VEGF-GFP fusion protein and confocal time-lapse microscopy, we observed the intracellular routing, secretion and immobilization of VEGF in scratch-activated living astrocytes. We found VEGF to be directly transported to cell-extracellular matrix attachments where it is incorporated into fibronectin fibrils. VEGF accumulated at ß1 integrin containing fibrillar adhesions and was translocated along the cell surface prior to internalization and degradation. We also found that only the astrocyte-derived, matrix-bound, and not soluble VEGF decreases ß1 integrin turnover in fibrillar adhesions. We suggest that polarized VEGF release and ECM remodeling by VEGF secreting cells is key to control the local concentration and signaling of VEGF. Our findings highlight the importance of astrocytes in directing VEGF functions and identify these mechanisms as promising target for angiogenic approaches.

Laser-Doppler microvascular measurements in the peri-implant areas of different osseointegrated bone conductor implant systems.

The objective of this study was to evaluate the impact of hydroxyapatite coating of newly designed osseointegrated fixtures' abutments on the postoperative complication rates. The integrity of peri-implant microcirculation was used as a marker to compare tissue viability after different surgical techniques. Laser-Doppler Flowmetry (LDF) measures alone, and coupled with heat provocation tests were applied to test the different microcircular patterns. Measures for 17 consecutively implanted patients (8 women, 9 men, ages ranged from 18 to 77 years) were recruited; seven with soft tissue reduction (STR); and 10 with soft tissue preservation (STP).Thirteen non-operated retro-auricular areas were examined as naive controls. In isotherm conditions the baseline blood flow remained stable in all groups. The naive control patients demonstrated significant changes of blood flux in the intact skin. The non-implanted yet previously operated contralateral sides of the patients demonstrated marginally lower (p = 0.09) blood flux index. The STR sides however, showed significantly lower (average 217 %) provoked blood flux compared to controls (p < 0.001). At the STP sides a maladaptation could be observed (average 316 %) compared to the contralateral sides (p = 0.53). STP sides demonstrated a significantly better blood flow improvement compared to the STR sides (p = 0.02). These results suggest a favorable postoperative condition of vascular microcirculation after STP, than after STR surgery. The possibly faster wound healing and lower potential complication rate may widen the inclusion criteria and maybe beneficial for the patient compliance with a better quality-of-life.

Dynamic connectivity among cortical layers in local and large-scale sensory processing.

Cortical processing of sensory stimuli typically recruits multiple areas, but how each area dynamically incorporates activity from other areas is not well understood. We investigated interactions between cortical columns of bilateral primary sensory regions (S1s) in rats by recording local field potentials and multi-unit activity simultaneously in both S1s with electrodes positioned at each cortical layer. Using dynamic connectivity analysis based on Granger-causal modeling, we found that, shortly after whisker stimulation (< 10 ms), contralateral S1 (cS1) already relays activity to granular and infragranular layers of S1 in the other hemisphere, after which cS1 shows a pattern of within-column interactions that directs activity upwards toward superficial layers. This pattern of predominant upward driving was also observed in S1 ipsilateral to stimulation, but at longer latencies. In addition, we found that interactions between the two S1s most strongly target granular and infragranular layers. Taken together, the results suggest a possible mechanism for how cortical columns integrate local and large-scale neocortical computation by relaying information from deeper layers to local processing in superficial layers.

Process of cortical network formation and impact of early brain damage.

PURPOSE OF REVIEW: The aim is to review mechanisms that are central to the formation of proper cortical circuitry and relevant to perinatal brain injury and premature birth. RECENT FINDINGS: Clinical investigations using noninvasive imaging techniques suggest that impaired connectivity of cortical circuitry is associated with perinatal adverse conditions. Recent experimental and translational studies revealed developmental mechanisms that are critical for circuit formation and potentially at risk in the perinatal period. These include existence of last wave genesis, migration and integration of gamma-aminobutyric acid (GABA) interneurons in the perinatal period; maturation of GABA interneuron networks that are central to critical period plasticity; transient connections by subplate neurons that guide thalamocortical connectivity, and a perineuronal microglia network that maintains axonal growth and neuronal survival as well as executing synaptic pruning. In addition, recent work has demonstrated that birth plays a key role in triggering the maturation cascade of cortical circuits. SUMMARY: Altered maturation of cortical circuits is an increasingly recognized aspect of perinatal injury and premature birth. Potential mechanisms are revealed but further translational studies are required to associate fine changes at the cellular and molecular level with imaging data in experimental models.

[New method for measuring the cortical auditory evoked potentials: the HEARLab]. / Új lehetoség az akusztikusan kiváltott corticalis válaszok mérésére: a HEARLab.

INTRODUCTION: Cortical auditory evoked potentials can provide objective information about the highest level of the auditory system. AIM: The purpose of the authors was to introduce a new tool, the "HEARLab" which can be routinely used in clinical practice for the measurement of the cortical auditory evoked potentials. In addition, they wanted to establish standards of the analyzed parameters in subjects with normal hearing. METHOD: 25 adults with normal hearing were tested with speech stimuli, and frequency specific examinations were performed utilizing pure tone stimuli. RESULTS: The findings regarding the latency and amplitude analyses of the evoked potentials confirm previously published results of this novel method. CONCLUSIONS: The HEARLAb can be a great help when performance of the conventional audiological examinations is complicated. The examination can be performed in uncooperative subjects even in the presence of hearing aids. The test is frequency specific and does not require anesthesia.

Resistance evolution to the first generation of genetically modified Diabrotica-active Bt-maize events by western corn rootworm: management and monitoring considerations.

Western corn rootworm (Diabrotica virgifera virgifera; WCR) is a major coleopteran maize pest in North America and the EU, and has traditionally been managed through crop rotation and broad-spectrum soil insecticides. Genetically modified Bt-maize offers an additional management tool for WCR and has been valuable in reducing insecticide use and increasing farm income. A concern is that the widespread, repeated, and exclusive deployment of the same Bt-maize transformation event will result in the rapid evolution of resistance in WCR. This publication explores the potential of WCR to evolve resistance to plant-produced Bt-toxins from the first generation of Diabrotica-active Bt-maize events (MON 863 and MON 88017, DAS-59122-7 and MIR604), and whether currently implemented risk management strategies to delay and monitor resistance evolution are appropriate. In twelve of the twelve artificial selection experiments reported, resistant WCR populations were yielded rapidly. Field-selected resistance of WCR to Cry3Bb1 is documented in some US maize growing areas, where an increasing number of cases of unexpected damage of WCR larvae to Bt-maize MON 88017 has been reported. Currently implemented insect resistance management measures for Bt-crops usually rely on the high dose/refuge (HDR) strategy. Evidence (including laboratory, greenhouse and field data) indicates that several conditions contributing to the success of the HDR strategy may not be met for the first generation of Bt-maize events and WCR: (1) the Bt-toxins are expressed heterogeneously at a low-to-moderate dose in roots; (2) resistance alleles may be present at a higher frequency than initially assumed; (3) WCR may mate in a non-random manner; (4) resistance traits could have non-recessive inheritance; and (5) fitness costs may not necessarily be associated with resistance evolution. However, caution must be exercised when extrapolating laboratory and greenhouse results to field conditions. Model predictions suggest that a 20 % refuge of non-Diabrotica-active Bt-maize can delay resistance evolution in WCR under certain conditions. This publication concludes that further research is needed to resolve the remaining scientific uncertainty related to the appropriateness of the HDR in delaying resistance evolution in WCR, resistance monitoring is essential to detect early warning signs indicating resistance evolution in the field, and that integrated pest management reliant on multiple tactics should be deployed to ensure effective long-term corn rootworm management and sustainable use of Bt-maize.