Header

UZH-Logo

Maintenance Infos

Two-photon imaging of neural activity and structural plasticity in the rodent spinal cord


Johannssen, H. Two-photon imaging of neural activity and structural plasticity in the rodent spinal cord. 2011, ETH Zurich, Faculty of Science.

Abstract

In my PhD thesis, I used two‐photon imaging to investigate neuronal circuits and
glia cells in the spinal cord of living mice. To achieve this, a major effort first was to
establish a mouse spinal cord preparation suitable for stable and long‐lasting
imaging experiments. Without adequate stabilisation, the spinal cord was prone to
large‐scale movement artefacts clearly hampering high‐resolution imaging in vivo.
To overcome these limitations, I employed strategies to optimise the animals
posture, namely rigid clamping of the vertebral column to isolate the spinal cord
from breathing‐related movements. In addition, I developed strategies to dampen
tissue movements remaining after posture optimisation. These improvements
made it possible to image the structural plasticity of genetically labelled microglia
cells with subcellular resolution for many hours in anesthetized mice. In a
paradigm of focal spinal cord injury, microglia became rapidly activated and
displayed high levels of filopodial motility clearly directed towards the injury site.
In addition, I adapted Ca2+ indicator loading techniques to stain neuronal networks
in the mouse superficial dorsal horn to visualize activity patterns of painprocessing
neurons. Despite the heavily myelinated surrounding tissue, neuronal
populations within the first two laminae could be visualized after Ca2+ indicator
loading. The preparation was sufficiently stable to for the first time resolve fast,
individual Ca2+ transients in the spinal cord of living rodents. In agreement with
the role of dorsal horn circuits in nociceptive processing I found low rates of
spontaneous activity but could reliably evoke increased activity levels by electrical
stimulation of primary afferent fibres in situ. Specifically, also natural sensory
stimulation applied to the paw elicited Ca2+ transients in a subset of dorsal horn
neurons.
In a parallel project, I collaborated with Klas Kullander’s group to investigate
activity patterns of identified Renshaw cells during an in vitro model of
locomotion. Using two‐photon Ca2+ imaging in the isolated neonatal mouse SC, we
found that several subclasses of Renshaw cells are differentially engaged in
ongoing locomotion. In addition, the activities of the different Renshaw cell populations were correlated with subgroups of simultaneously recorded
motoneurons. Afferent inputs delivered during ongoing locomotion perturbed the
locomotor rhythm and the nature of perturbations depended on stimulus timing
during either the flexor‐ or extensor‐related cycle phase. On the local circuit level,
we observed that correlations between specific Renshaw cells and motoneuron
subpopulations were boosted by sensory input and that this effect also depended
on stimulus timing. In a broader context, these results can be interpreted as
reflections of synaptic strengthening of developing locomotor modules by sensory
inputs.

Abstract

In my PhD thesis, I used two‐photon imaging to investigate neuronal circuits and
glia cells in the spinal cord of living mice. To achieve this, a major effort first was to
establish a mouse spinal cord preparation suitable for stable and long‐lasting
imaging experiments. Without adequate stabilisation, the spinal cord was prone to
large‐scale movement artefacts clearly hampering high‐resolution imaging in vivo.
To overcome these limitations, I employed strategies to optimise the animals
posture, namely rigid clamping of the vertebral column to isolate the spinal cord
from breathing‐related movements. In addition, I developed strategies to dampen
tissue movements remaining after posture optimisation. These improvements
made it possible to image the structural plasticity of genetically labelled microglia
cells with subcellular resolution for many hours in anesthetized mice. In a
paradigm of focal spinal cord injury, microglia became rapidly activated and
displayed high levels of filopodial motility clearly directed towards the injury site.
In addition, I adapted Ca2+ indicator loading techniques to stain neuronal networks
in the mouse superficial dorsal horn to visualize activity patterns of painprocessing
neurons. Despite the heavily myelinated surrounding tissue, neuronal
populations within the first two laminae could be visualized after Ca2+ indicator
loading. The preparation was sufficiently stable to for the first time resolve fast,
individual Ca2+ transients in the spinal cord of living rodents. In agreement with
the role of dorsal horn circuits in nociceptive processing I found low rates of
spontaneous activity but could reliably evoke increased activity levels by electrical
stimulation of primary afferent fibres in situ. Specifically, also natural sensory
stimulation applied to the paw elicited Ca2+ transients in a subset of dorsal horn
neurons.
In a parallel project, I collaborated with Klas Kullander’s group to investigate
activity patterns of identified Renshaw cells during an in vitro model of
locomotion. Using two‐photon Ca2+ imaging in the isolated neonatal mouse SC, we
found that several subclasses of Renshaw cells are differentially engaged in
ongoing locomotion. In addition, the activities of the different Renshaw cell populations were correlated with subgroups of simultaneously recorded
motoneurons. Afferent inputs delivered during ongoing locomotion perturbed the
locomotor rhythm and the nature of perturbations depended on stimulus timing
during either the flexor‐ or extensor‐related cycle phase. On the local circuit level,
we observed that correlations between specific Renshaw cells and motoneuron
subpopulations were boosted by sensory input and that this effect also depended
on stimulus timing. In a broader context, these results can be interpreted as
reflections of synaptic strengthening of developing locomotor modules by sensory
inputs.

Statistics

Altmetrics

Downloads

93 downloads since deposited on 17 Jan 2012
15 downloads since 12 months
Detailed statistics

Additional indexing

Item Type:Dissertation
Referees:Helmchen F, Schwab M E, Zeilhofer H U
Communities & Collections:04 Faculty of Medicine > Brain Research Institute
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:2011
Deposited On:17 Jan 2012 19:37
Last Modified:12 Aug 2017 21:46
Additional Information:Diss., Eidgenössische Technische Hochschule ETH Zürich, Nr. 19666, 2011; Die Arbeit wurde erstellt am Institut für Hirnforschung der Univ. Zürich.
Free access at:Related URL. An embargo period may apply.
Publisher DOI:https://doi.org/10.3929/ethz-a-006716087
Related URLs:http://opac.nebis.ch/F/?local_base=NEBIS&CON_LNG=GER&func=find-b&find_code=SYS&request=006716087

Download