Jeroen Aerts

The cross-modal aspect of mouse visual cortex plasticity induced by monocular enucleation is age dependent

Monocular enucleation (ME) drastically affects the contralateral visual cortex, where plasticity phenomena drive specific adaptations to compensate for the unilateral loss of vision. In adult mice, complete reactivation of deprived visual cortex involves an early visually driven recovery followed by multimodal plasticity 3 to 7 weeks post ME (Van Brussel et al. [ ] Cereb. Cortex 21:2133–2146). Here, we specifically investigated the age dependence of the onset and the exact timing of both ME‐induced reactivation processes by comparing cortical activity patterns of mice enucleated at postnatal day (P) 45, 90, or 120. A swifter open‐eye potentiated reactivation characterized the binocular visual cortex of P45 mice. Nevertheless, even after 7 weeks, the reactivation remained incomplete, especially in the monocular cortex medial to V1. In comparison with P45, emergent cross‐modal participation was demonstrated in P90 animals, although robust reactivation similar to enucleated adults (P120) was not achieved yet. Concomitantly, at 7 weeks post ME, somatosensory and auditory cortex shifted from a hypoactive state in P45 to hyperactivity in P120. Thus, we provide evidence for a presensitive period in which gradual recruitment of cross‐modal recovery upon long‐term ME coincides with the transition from adolescence to adulthood in mice. J. Comp. Neurol. 522:950–970, 2014.

Ocular Hypertension Results in Retinotopic Alterations in the Visual Cortex of Adult Mice

Abstract Purpose: Glaucoma is a group of optic neuropathies characterized by the loss of retinal ganglion cells (RGCs). Since ocular hypertension (OHT) is a main risk factor, current therapies are predominantly based on lowering eye pressure. However, a subset of treated patients continues to lose vision. More research into pathological mechanisms underlying glaucoma is therefore warranted in order to develop novel therapeutic strategies. In this study we investigated the impact of OHT from eye to brain in mice. Methods: Monocular hypertension (mOHT) was induced in CD-1 mice by laser photocoagulation (LP) of the perilimbal and episcleral veins. The impact on the retina and its main direct target area, the superficial superior colliculus (sSC), was examined via immunostainings for Brn3a, VGluT2 and GFAP. Alterations in neuronal activity in V1 and extrastriate areas V2L and V2M were assessed using in situ hybridization for the activity reporter gene zif268. Results: Transient mOHT resulted in diffuse and sectorial RGC degeneration. In the sSC contralateral to the OHT eye, a decrease in VGluT2 immunopositive synaptic connections was detected one week post LP, which appeared to be retinotopically linked to the sectorial RGC degeneration patterns. In parallel, hypoactivity was discerned in contralateral retinotopic projection zones in V1 and V2. Despite complete cortical reactivation 4 weeks post LP, in the sSC no evidence for recovery of RGC synapse density was found and also the concomitant inflammation was not completely resolved. Nevertheless, sSC neurons appeared healthy upon histological inspection and subsequent analysis of cell density revealed no differences between the ipsi- and contralateral sSC. Conclusion: In addition to RGC death, OHT induces loss of synaptic connections and neuronal activity in the visual pathway and is accompanied by an extensive immune response. Our findings stress the importance of looking beyond the eye and including the whole visual system in glaucoma research.

Development and Validation of a Small Single-domain Antibody That Effectively Inhibits Matrix Metalloproteinase 8

A detrimental role for matrix metalloproteinase 8 (MMP8) has been identified in several pathological conditions, e.g., lethal hepatitis and the systemic inflammatory response syndrome. Since matrix MMP8-deficient mice are protected in the above-mentioned diseases, specific MMP8 inhibitors could be of clinical value. However, targeting a specific matrix metalloproteinase remains challenging due to the strong structural homology of matrix metalloproteinases, which form a family of 25 members in mammals. Single-domain antibodies, called nanobodies, offer a range of possibilities toward therapy since they are easy to generate, express, produce, and modify, e.g., by linkage to nanobodies directed against other target molecules. Hence, we generated small MMP8-binding nanobodies, and established a proof-of-principle for developing nanobodies that inhibit matrix metalloproteinase activity. Also, we demonstrated for the first time the possibility of expressing nanobodies systemically by in vivo electroporation of the muscle and its relevance as a potential therapy in inflammatory diseases.

Synthesis and Validation of a Hydroxypyrone-Based, Potent, and Specific Matrix Metalloproteinase-12 Inhibitor with Anti-Inflammatory Activity In Vitro and In Vivo

A hydroxypyrone-based matrix metalloproteinase (MMP) inhibitor was synthesized and assayed for its inhibitory capacity towards a panel of ten different MMPs. The compound exhibited selective inhibition towards MMP-12. The effects of inhibition of MMP-12 on endotoxemia and inflammation-induced blood-cerebrospinal fluid barrier (BCSFB) disruption were assessed both in vitro and in vivo. Similar to MMP-12 deficient mice, inhibitor-treated mice displayed significantly lower lipopolysaccharide- (LPS-) induced lethality compared to vehicle treated controls. Following LPS injection Mmp-12 mRNA expression was massively upregulated in choroid plexus tissue and a concomitant increase in BCSFB permeability was observed, which was restricted in inhibitor-treated mice. Moreover, an LPS-induced decrease in tight junction permeability of primary choroid plexus epithelial cells was attenuated by inhibitor application in vitro. Taken together, this hydroxypyrone-based inhibitor is selective towards MMP-12 and displays anti-inflammatory activity in vitro and in vivo.

A highly reproducible and straightforward method to perform in vivo ocular enucleation in the mouse after eye opening.

Enucleation or the surgical removal of an eye can generally be considered as a model for nerve deafferentation. It provides a valuable tool to study the different aspects of visual, cross-modal and developmental plasticity along the mammalian visual system1-4. Here, we demonstrate an elegant and straightforward technique for the removal of one or both eyes in the mouse, which is validated in mice of 20 days old up to adults. Briefly, a disinfected curved forceps is used to clamp the optic nerve behind the eye. Subsequently, circular movements are performed to constrict the optic nerve and remove the eyeball. The advantages of this technique are high reproducibility, minimal to no bleeding, rapid post-operative recovery and a very low learning threshold for the experimenter. Hence, a large amount of animals can be manipulated and processed with minimal amount of effort. The nature of the technique may induce slight damage to the retina during the procedure. This side effect makes this method less suitable as compared to Mahajan et al. (2011)5 if the goal is to collect and analyze retinal tissue. Also, our method is limited to post-eye opening ages (mouse: P10 - 13 onwards) since the eyeball needs to be displaced from the socket without removing the eyelids. The in vivo enucleation technique described in this manuscript has recently been successfully applied with minor modifications in rats and appears useful to study the afferent visual pathway of rodents in general.

Effects of medium renewal and handling stress on life history traits in Daphnia

The zooplankton genus Daphnia is used as a model organism in ecological, ecotoxicological and evolutionary research. It is often used in experimental laboratory setups to examine life history traits under a variety of factors inducing stress. One type of stress, handling stress because of manipulation during the experiments, however, is often neglected. In a standard life history experiment, we found that several kinds of experimental manipulation may have a significant impact on the life history response of test animals. Especially renewal of the medium may reduce the reproductive output of the individuals significantly. In every experiment where the reproduction of Daphnia is an important trait under study, the negative effects of medium renewal and handling stress should be considered and care should be taken to keep handling of the individuals to a minimum.

MS imaging and mass spectrometric synaptosome profiling identify PEP-19/pcp4 as a synaptic molecule involved in spatial learning in mice

The Morris water maze (MWM) spatial learning task has been demonstrated to involve a cognitive switch of action control to serve the transition from an early towards a late learning phase. However, the molecular mechanisms governing this switch are largely unknown. We employed MALDI MS imaging (MSI) to screen for changes in expression of small proteins in brain structures implicated in the different learning phases. We compared mice trained for 3days and 30days in the MWM, reflecting an early and a late learning phase in relation to the acquisition of a spatial learning task. An ion with m/z of 6724, identified as PEP-19/pcp4 by top-down tandem MS, was detected at higher intensity in the dorsal striatum of the late learning phase group compared with the early learning phase group. In addition, mass spectrometric analysis of synaptosomes confirmed the presence of PEP-19/pcp4 at the synapse. PEP-19/pcp4 has previously been identified as a critical determinant of synaptic plasticity in locomotor learning. Our findings extend PEP-19/pcp4 function to spatial learning in the forebrain and put MSI forward as a valid and unbiased research strategy for the discovery and identification of the molecular machinery involved in learning, memory and synaptic plasticity. This article is part of a Special Issue entitled: MALDI Imaging, edited by Dr. Corinna Henkel and Prof. Peter Hoffmann.