Abstract
In ventral mesencephalic organotypic tissue cultures, two timely separated sequences of nerve fiber growth have been observed. The first appearing nerve fiber pattern is a long-distance outgrowth that occurs before astrocytes start to proliferate and migrate to form an astrocytic monolayer that finally surrounds the tissue slice. These long-distance growing nerve fibers are retracted as the astrocytes migrate, and are followed by a secondary outgrowth. The secondary outgrowth is persistent in time but reaches short distances, comparable with outgrowth seen from a dopaminergic graft implanted to the brain. The present study was focused on the interaction between the astrocytes and the long-distance growing non-glial associated nerve fibers. Cross talk between astroglia and neurite formation might occur through the integrin-associated protein CD47. CD47 serves as a ligand for signal regulatory protein (SIRP) α and as a receptor for the extracellular matrix protein thrombospondin-1 (TSP-1). Embryonic day 14 ventral mesencephalic tissue from CD47+/+ and CD47−/− mice was used to investigate astrocytic migration and the tyrosine hydroxylase (TH) –positive outgrowth that occurred remote from the astrocytes. TH-immunohistochemistry demonstrated that the non-glial-associated nerve fiber outgrowth in CD47−/− cultures reached significantly longer distances and higher density compared to nerve fibers formed in CD47+/+ cultures at 14 days in vitro. These nerve fibers often had a dotted appearance in CD47+/+ cultures. No difference in the astrocytic migration was observed. Further investigations revealed that the presence of CD47 in control culture did neither hamper non-glial-associated growth through SIRPα nor through TSP-1 since similar outgrowth was found in SIRPα mutant cultures and in CD47+/+ cultures treated with blocking antibodies against the TSP-1, respectively, as in the control cultures. In conclusion, long-distance growing nerve fiber formation is promoted by the absence of CD47, even though the presence of astrocytes is not inhibited.
Highlights
Regeneration in the adult brain involves neurotrophic factors, neuroinhibitors, cell adhesion, and extracellular matrix molecules that may affect the regenerative process
The results revealed that the distance for tyrosine hydroxylase (TH)-positive nerve fiber outgrowth was affected by time without any significance in interaction between time and genotypes
The total distance reached by the nerve fiber outgrowth was around 1 mm at 7 days in vitro (DIV) and around 3.5 mm at 14 DIV for CD472/2 cultures, which was longer than the astrocytic migration had reached at the respective time points (Fig. 2c)
Summary
Regeneration in the adult brain involves neurotrophic factors, neuroinhibitors, cell adhesion, and extracellular matrix molecules that may affect the regenerative process These molecules are often produced by the glial cells and affects the nerve regeneration for instance in the spinal cord after injury, where regeneration beyond the glial scar is problematic. Much effort has been dedicated to study reinnervation of the striatum induced by transplanted dopaminergic cells in animal models of Parkinson’s disease [1,2] In this situation no strong correlation to inhibited graft outgrowth has been accounted to the astrocytic influence, still graft outgrowth is limited to small zones surrounding the transplants, which limits the effect of the transplant, unless multiple graft sites are made to cover most of the dopamine-denervated portion of the striatum [3]. To investigate what influences nerve fiber growth and what may make the graft outgrowth to halt at a certain distance from the grafted tissue, organotypic tissue cultures have been employed
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