Institutionen för neurovetenskap / Department of Neuroscience
Abstract
Spinal motoneurons integrate a vast synaptic input and are the final
conveyors of motor commands to skeletal muscle. Motoneuron regeneration
includes altered contacts between the motoneuron and several other cell
types. The severed axon elongates along Schwann cells in the nerve, while
other types of glia interact closely with lesioned motoneurons in the
spinal cord. Simultaneously, cell contacts are disrupted when synapses
are lost from lesioned motoneurons in a process called synaptic
stripping.
The aim of this thesis project was to identify potential mediators of
cell interactions during motoneuron regeneration, with special emphasis
on molecules that could be of importance for synaptic remodeling. A
reduced expression of cell adhesion molecules has been speculated to be
involved in synaptic stripping of motoneurons. Over the last years,
several such molecules have been described which influence formation and
maintenance of synapses, and also mediate adhesion in other events that
occur during motoneuron regeneration, such as axon guidance and
myelination. We demonstrate that peripheral axotomy of sciatic
motoneurons results in altered expression of several cell adhesion
molecules, many of which are previously unstudied in this context. Some
of these changes indicate possible involvement of the molecules in
synapse elimination, whereas other molecules may be involved in other
regenerative events.
Specifically, the expression of nectin-1 and -3 increased in lesioned
motoneurons, as did the expression of nectin-3-binding necl-5. Nectin
proteins did not localize to synapses on spinal motoneurons, but instead
to neuronal processes and glia, both within the spinal cord and in the
lesioned sciatic nerve. Immunoreactivity for N-cadherin localized to
synapses on the surface of motoneurons and was reduced after sciatic
nerve transection (SNT). Motoneuron expression of mRNA encoding
N-cadherin was not altered after axotomy, and immunoreactivity for the
molecule increased in the severed nerve. Axotomy also resulted in altered
expression of SynCAM3/necl-1 and SynCAM4/necl-4 in the nerve, which
indicates possible involvement of these molecules in remyelination.
SynCAM1/necl-2, SynCAM2/necl-3, and neuroligin (NLG) -2 and -3 were
expressed by unlesioned motoneurons, and SynCAM immunoreactivity
localized to synapses on motoneuron cell bodies. In vitro these molecules
have synapse-inducing properties, and following SNT, expression of
SynCAM1 and NLG2 and -3 decreased rapidly, prior to loss of staining for
synaptophysin in the motoneuron pool. SynCAM1 expression correlated to
loss and return of synapses in regeneration after SNT. NLG expression
decreased to a smaller degree after sciatic nerve crush than after SNT,
although the loss of synapses was similar in both lesion models. Finally,
while this work was ongoing, complement-tagging of CNS synapses for
removal was demonstrated to occur in the visual system. We investigated
whether complement could be involved also in synapse removal from
axotomized motoneurons. Complement C3-/- mice displayed reduced synaptic
stripping after lesion, a larger upregulation of growth-associated
protein 43 in motoneurons, and a more rapid restoration of motor
function.
We conclude that the motoneuron response to axotomy involves
downregulation of several synaptic adhesion molecules. Expression of
SynCAM1 correlates closely to the loss and return of synapses but the
magnitude of the downregulation of NLGs does not seem to reflect the
magnitude of the loss of synapses. Contact with the distal nerve stump
may stimulate expression of NLGs, but does not seem to influence that of
SynCAM1. Expression of NLGs and SynCAM1 does not seem to be the sole
determinant of the elimination of synapses, since the expression pattern
of these molecules was similar in mice with altered synaptic stripping
and wild type mice. We also conclude that complement C3 is required for
normal synapse elimination. Thus, complement may be a potential target in
therapeutic attempts to preserve synaptic circuits
