Book of abstracts - British Neuroscience Association

4.13
Regulation of Egr1 and Egr3 following the induction of LTP in
CA1 of the hippocampus.
Cheval H, Laroche S, Davis S
NAMC, UMR 8620, CNRS, Univ Paris-Sud, 91405 Orsay, France
The gene encoding Egr1 (zif268) is regulated and necessary for late
phases of LTP in dentate gyrus and the consolidation of a number of
different forms of memory. Very little data exists about the potential
role of Egr1 and other members of this family of transcription factors in
plasticity in CA1. We have examined regulation of both Egr1 and Egr3
following the induction of LTP in CA1 in both rats and Egr1-/- and +/+
mice. Preliminary evidence shows LTP induced in +/+ mice is
associated with an increase in Egr1 protein levels. However,
regulation of the protein is not specific to LTP as pseudotetanus also
induces an increase in Egr1 protein level. In mutant mice, LTP is
attenuated and western blotting confirms the lack of protein,
suggesting that Egr1 is neither necessary nor specific to LTP in CA1.
Induction of LTP also induced a similar pattern of overexpression of
Egr3 in both wildtype and mutant mice with regulation of the protein
restricted to the Ą isoform, suggesting that Egr3 regulation may be
linked with regulation of Egr1. As a transcription factor, Egr1/Egr3
presumably bind to downstream gene targets with a consensus ERE
sequence on their promoter. We are conducting EMSA and supershift
assays in rat to determine whether Egr1 and Egr3 functional binding to
promoter regions sequences is increased following induction of LTP.
Preliminary evidence suggests this to be the case and supershift
experiments will be conduced to examine the specificity of Egr1 and
Egr3 binding to the ERE sequence.
4.14
Multielectrode array-based platform for analysis of the synaptic
transmission and plasticity phenotypes in mutant mice
Kopanitsa MV, Afinowi NO, Grant SGN
Genes to Cognition Programme, The Wellcome Trust Sanger Institute,
Hinxton, Cambridge CB10 1SA
The Genes to Cognition Programme (G2C, www.genes2cognition.org) is a
multidisciplinary initiative to study genes involved in brain functions and
behaviour. An important part of G2C is production and characterisation of
transgenic mice. We sought to employ multielectrode array (MEA)
technology for the high-throughput electrophysiological assessment of
mutant animals. We have shown previously (Kopanitsa et al., BMC
Neuroscience 7: 61) that MEAs allow for efficient recording of synaptic
potentials and facilitate LTP studies in hippocampal slices.
To further validate MEA-based approaches, we studied several mutants
where deficiencies in LTP had been documented. Synaptic responses were
recorded in the CA1 area of hippocampal slices upon stimulation of
Schäffer collaterals. LTP was induced in one of the two stimulated
pathways by high frequency tetani and/or by theta-burst stimulation. We
have observed deficient LTP phenotypes in GluR-A -/-; SynGAP +/- and
NR2A ΔC/ΔC mice, which was consistent with published reports. Also, we
found a decrease of LTP in the NR2B/ΔC mice, which suggested that
integrity of cytoplasmic tails of both NR2A and NR2B subunits is important
for plasticity. The MEA-based platform was also used to investigate
synaptic transmission and LTP in a number of novel strains created in our
lab. We conclude that MEA technology is a rapid and efficient platform for
analysis of synaptic transmission and plasticity in mutant mice. In addition
to the throughput, an advantage of MEAs is that they allow standardising
recording conditions during LTP experiment, which is extremely important
for comparing the severity of different mutant phenotypes.
5.01
Human bone marrow stromal cells promote nerve growth over
the major nerve-inhibitory molecules found in the injured spinal
cord.
Wright K T, El Masri W, Osman A, Roberts S, Chamberlain G, Ashton
BA, Johnson WE
Centre for Spinal Studies, Robert Jones and Agnes Hunt Orthopaedic
Hospital, Oswestry, Shropshire SY10 7AG, UK; Institute for Science
and Technology in Medicine, Keele University, Keele, Staffordshire
ST5 5BG, UK.
Chondroitin sulphated proteoglycans (CSPG), myelin associated
glycoprotein (MAG) and Nogo inhibit nerve growth in vivo. Their
inhibitory influence in spinal cord injury (SCI) has been shown in
animal models wherein treatments that reduced their presence,
integrity or blocked their activity promoted axonal regeneration and
functional recovery. Transplantation of bone marrow stromal cells
(MSC) into the injured spinal cord induced similar results, but it is still
unclear how the improvements noted in these animals were achieved.
We have examined the potential for human MSC isolated from SCI
patients to promote nerve growth across inhibitory molecules using an
established in vitro model of nerve growth. As previously described
CSPG, MAG and Nogo were each found to inhibit neurite outgrowth in
a concentration dependant manner, this effect was diminished in the
presence of MSC. Time-lapse video microscopy demonstrated that
MSC acted as “cellular bridges” and also “towed” neurites over
inhibitory substrates. Whereas conditioned medium from MSC cultures
stimulated neurite outgrowth over type I collagen, it did not promote
outgrowth over CSPG, MAG or Nogo.
5.02
Mesenchymal stem cell- derived soluble factors and their instructive
effects on neural stem and progenitor cells
Hardy S A, Croft A P, Przyborski S A
1School of Biological and Biomedical Sciences, University of Durham,
South Road, Durham, DH1 3LE; 2ReInnervate Ltd, Durham, DH1 3HP
Mesenchymal stem cell (MSC) transplantation in animal models of
neurological disease has resulted in neurological improvement, however,
the mechanism by which this occurs is under debate. Evidence suggests
that MSCs have neurogenic potential and undergo differentiation into
neural tissue to replace cells damaged in disease (trans-differentiation).
These data are conflicting as others argue that MSCs do not transdifferentate
but instead fuse with host cells adopting their phenotype.
However, both mechanisms are unlikely to fully account for the
improvements seen. We are currently investigating a third mechanism
whereby transplanted MSCs promote endogenous repair of neurologically
damaged areas by the release of trophic factors and cytokines e.g. NGF
and BDNF. We have shown that MSCs produce factors that instruct neural
stem cells (NSCs) to adopt neuronal and glial lineages, and this is
dependent on the developmental status of the MSC. Under standard
culture conditions MSCs are negative for neural antigens, and produce
factors which instruct NSCs to adopt a predominantly astrocytic fate.
Conversely, we have shown that MSCs induced to form neurosphere-like
structures, which are positive for neural antigens such as nestin and GFAP,
instruct NSCs to adopt a predominantly neuronal fate. Such events are
likely to contribute to the functional improvements observed in disease
models following MSC transplantation. We are currently in the preliminary
stages of identifying these unknown factors.
These findings suggest that MSC transplantation may promote axonal
regeneration by stimulating nerve growth via secreted factors and also
by reducing the nerve-inhibitory effects of extracellular molecules.
Further experimentation using this model will help identify how MSC
are able to migrate over nerve-inhibitory molecules and how they
stimulate co-localised nerve growth. This thereby may permit
appropriate molecular targeting to further enhance nerve growth in
vivo.
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