Background: Neurons fail to be replaced endogenously, but transplantation provides a feasible option for replacement, if neurons integrate adequately.
Hypothesis: In the previous funding period we showed that the injury condition and age profoundly influence the input connectome and we discovered a severe limitation of output connectivity due to the lack of myelination, i.e., critical checkpoints for integration mediated by glia-neuron interactions.
Strategy: To optimize the input and output connectome we utilize a candidate approach as well as unbiased RABID-seq to determine the interactions of transplanted Neurons (tN) with their glial neighbors regulating the input synaptic connections and myelination.
Summary
Neurons are lost in many disease conditions and fail to be replaced endogenously. However, transplantation of neurons is a clinically relevant approach to replace neurons if they adequately integrate into the circuitry. Here we explore the neuron-glia interactions as critical checkpoints for adequate inputs to and outputs of transplanted neurons (tNs). In the previous funding period, we focused on the input connectome in different disease conditions. We observed a particularly high number of input neurons as monitored by monosynaptic Rabies virus tracing in aging and amyloid loaded brains (Thomaset al., 2022) and transiently in brains with traumatic brain injury (Grade et al., 2022). To understand and improve this loss of input, synapses at morphological, light-microscopic and ultrastructural level as well as by functional electrophysiology were examined. These data show no loss, but rather maturation of spines. However, postsynaptic potentials become less frequent over time, suggesting loss, or silencing of synapses. Ultrastructural analysis shows many synapses on shafts of the tN which could be subject to pruning. To understand the molecular mechanisms of synapse reduction we examined the input connectome of tNs in C1qa- and TREM2 knock-out mice that show opposite phenotypes with excessive inputs in C1qa KO brains at early time points followed by a reduction of input neurons, while excessive input neurons are found mostly at later stages after transplantation in TREM2 KO brains. This suggests a role of TREM2 in restricting synapse loss at later stages and C1qa inhibiting establishment of initial synaptic contacts. Here we aim to identify the molecular mechanisms mediating this checkpoint using RABID-seq to explore receptor ligand pairs between tNs and interacting glia at the single cell level. We will also explore the functional aspects of these initially or later increased synaptic contacts to achieve the final aim of a correct input connectivity. To probe the adequate output of tNs, we examined their axonal myelination and found virtually none of the axons of tNs to be myelinated, while endogenous neurons labelled in the same manner, both at light microscopic and ultrastructural level, were myelinated. Here we will team up with the experts in myelination within our CRC to probe for missing receptor-ligand interactions between tNs and oligodendrocyte progenitors that should also emerge from the RABID-seq approach, as well as utilize chemogenic tools in collaboration in the CRC to activate the tNs. In a further CRC collaboration, we will employ pharmacological treatments used in demyelinating disease to enhance the myelination of tN axons. Ultimately these projects aim to achieve adequate input and output and thus correct circuit integration of tNs also in disease conditions.
