Immune recognition molecules in synaptic plasticity and regeneration of spinal motoneurons

Abstract

This thesis is based on the emerging concept that pattern recognition molecules, originally characterized in the immune system, may be expressed and used by neurons to mediate non-immune functions. In line with this concept, the major histocompatibility complex (MHC) class I and certain complements proteins have been implicated in synaptic plasticity in the developing visual system.

In Paper I, we studied the expression of MHC class I mRNAs and proteins in normal and axotomized spinal mouse motoneurons. Two mRNAs encoding classical MHC class I molecules (H2-Kb and H2-Db) were moderately expressed in uninjured motoneuron cell bodies. After a peripheral nerve lesion, both mRNAs were strongly up-regulated by axotomized motoneurons and surrounding glial cells. Using a MHC class I antibody with affinity for H2-Db, we observed moderate immunoreactivity (IR) in the cell bodies of a subpopulation of uninjured spinal motoneurons. After a peripheral nerve lesion, H2-Db IR was strongly increased in activated microglia. In contrast to the in situ hybridization results, the H2-Db IR remained unchanged in axotomized motoneuron cell bodies. We then further investigated the in vivo motoneuron expression of H2-Db in the periphery. H2-Db IR was detected in a subpopulation of axons in the sciatic nerve and at the presynaptic side of the neuromuscular junction (NMJ) in hind limb muscles. When studying mice deficient in classical MHC class I (Kb-/-Db-/-), we observed abnormal dynamic changes in NMJ density during muscle reinnervation and delayed motor recovery after a sciatic nerve crush (SNC). During the reinnervation phase, Kb-/-Db-/- mice also displayed an attenuated proliferation of terminal Schwann cells at NMJs compared to wild-type mice (WT). Interestingly, we found expression of the paired immunoglobulin receptor B in dissociated Schwann cells and histological sections from the sciatic nerve.

In order to investigate the role of MHC class I proteins in central motoneuron plasticity, we studied synaptic elimination from axotomized motoneuron cell bodies at the ultrastructural level in Paper II. In contrast to a previous publication by Shatz et al. 2000, axotomized motoneurons in mice lacking functional MHC class I (TAP1-/- and beta2m-/-) displayed an increased synaptic elimination compared to WT mice. Moreover, in beta2m-/- mice the remaining terminals were randomly dispersed along the cytoplasmic membrane in difference to WT animals where they were tightly clustered. When analyzing the types of synaptic terminals that were retracted in the beta2m-/- mice, we found a preferential loss of inhibitory terminals. In parallel, axonal regeneration appeared to be hampered in the absence of functional MHC class I molecules.

Since complement-deficient animals (C1q-/- and C3-/-) are shown to display a phenotype resembling that of MHC class I-deficient mice regarding synaptic plasticity in the visual system, we investigated the role for complement proteins in adult motoneuron plasticity in Paper III. In accordance with a previous study by Steven et al. 2007, C3-/- deficient animals displayed a diminished reduction in synapse density and covering of axotomized motoneurons. The histological expression pattern of C1q and C3 in the spinal cord was somewhat hard to interpret. We found a clear up-regulation of complement mRNA and protein in the axotomized sciatic motor pool, but we have so far failed to determine the subcellular localization with certainty. Nonetheless, we found complement IR in close association with the motoneuron surface and with presynaptic terminals on proximal dendrites and with surrounding glial cells. In addition, C3-/- animals recovered motor function more rapidly after a SNC.

In conclusion, we have investigated and found evidence of new roles for classical immune molecules in motoneurons with regard to synaptic plasticity and regeneration. The subcellular expression and signaling pathways remain to be described before specific functions and sites of action for these molecules can be determined. Further studies of neuronal immune molecules will be important in order to gain insight into the mechanisms of cellular interaction between different types of neurons or glial cells.