A Nogo-Like Signaling Perspective from Birth to Adulthood and in Old Age: Brain Expression Patterns of Ligands, Receptors and Modulators

Abstract

An appropriate strength of Nogo-like signaling is important to maintain synaptic homeostasis in the CNS. Disturbances have been associated with schizophrenia, MS and other diseases. Blocking Nogo-like signaling may improve recovery after spinal cord injury, stroke and traumatic brain injury. To understand the interacting roles of an increasing number of ligands, receptors and modulators engaged in Nogo-like signaling, the transcriptional activity of these genes in the same brain areas from birth to old age in the normal brain is needed. Thus, we have quantitatively mapped the innate expression of 11 important genes engaged in Nogo-like signaling. Using in situ hybridization, we located and measured the amount of mRNA encoding Nogo-A, OMgp, NgR1, NgR2, NgR3, Lingo-1, Troy, Olfactomedin, LgI1, ADAM22, and MAG, in 18 different brain areas at six different ages (P0, 1, 2, 4, 14, and 104 weeks). We show gene- and area-specific activities and how the genes undergo dynamic regulation during postnatal development and become stable during adulthood. Hippocampal areas underwent the largest changes over time. We only found differences between individual cortical areas in Troy and MAG. Subcortical areas presented the largest inter-regional differences; lateral and basolateral amygdala had markedly higher expression than other subcortical areas. The widespread differences and unique expression patterns of the different genes involved in Nogo-like signaling suggest that the functional complexes could look vastly different in different areas.

Keywords: Lingo-1; NgR1; Nogo-A; OMgp; Olfactomedin; Troy; development; hippocampus.

Figure 1

A schematic representation of the Nogo system. Ligands: Nogo-A, OMgp, MAG, Blys. Receptors: NgR1, NgR2, NgR3, PirB, ADAM22, S1PR2. Coreceptors: Troy, P75, Lingo-1, Amigo3. Modulators: LOTUS, LgI1, Olfactomedin, CSPG, MT3-MMP. Picture from Karlsson et al. (2017).

Figure 2

Oligonucleotide DNA probes.

Figure 3

List over regions and abbreviations.

Figure 4

High resolution scans from X-ray film autoradiographs. Representative autoradiograms from in situ hybridization performed with ³³P-labeled oligonucleotides for four different genes (A: Olfactomedin 1 mRNA 1 week; B: Nogo-A mRNA 2 weeks; C: Lingo-1 mRNA 4 weeks and D: NgR1 mRNA 104 weeks). These scans were used for measurements of gene expression in the 18 different regions (Figure 3).

Figure 5

Pooled average expression of all subregions in the three main regions over time for each gene; (A) cortex, (B) hippocampal formation and (C) subcortical structures. Subregions are found in Figure 3. Y-axis: expression in nCi/g.

Figure 6

Distribution of 175 significant changes (p < 0.05) of mRNA expression over time for the 11 studied Nogo-related genes in the three main regions; cortex, hippocampal formation and subcortical areas. The changes are later described and visualized in charts in the paragraphs for each gene. In this table, significant change of mRNA expression between either neighboring time points (1 week – P0, 2 weeks – 1 week or 4 weeks – 2 weeks) or when comparing the mRNA expression in the adult 14 weeks old mice to that in P0, 1 or 2 weeks old mice, is coded using a green scale concerning increases, and decreases using a red scale. Notably, all genes except MAG decrease with time or lack a significant change in cortical areas, while hippocampal levels mostly increase with time. It is also the hippocampal formation that had significant changes in all but 2 of the investigated genes. Changes were detected with the help of a mixed model analysis, see statistics section.

Figure 7

Distribution of 339 significant differences (p < 0.05) between regions for the 11 studied Nogo-related genes in cortex, hippocampal formation and subcortical areas. In the cortex, most of the genes had similar expression in most cortical areas investigated. In the hippocampus, 7/10 genes were differently expressed in the different regions while 3 had an even expression. Inter-regional mRNA expression level differences were foremost found in the heteregenous subcortical regions.

Figure 8

Distribution of significant changes (p < 0.05) of mRNA levels per region during maturation and aging. During maturation, most changes were observed between week 2 and 4. When comparing the mRNA expression levels in 14 weeks old mice to the young ages it becomes clear that the system gradually reaches its adult expression level between 2 and 4 weeks and remains remarkably stable into aging.

Figure 9

NgR1 (A–C), Lingo-1 (E–G) and Troy (I–K) mRNA levels in 18 brain regions over time. The x-axis indicates age in weeks, the y-axis indicates levels of mRNA expression in nCi/g, and is individual for each gene and region. Significant changes defined as p < 0,05 are mentioned in the text for each gene. Heatmaps of the three genes (D,H,L) visualizing expression in 14 week old mice.

Figure 10

Heatmap of Nogo-A sections at P0, 1 and 2 weeks, demonstrating a global decline of mRNA expression during postnatal development.

Figure 11

Graphs showing expression levels of mRNA encoding Nogo-A (A–C) and OMgp (E–G) in 18 cerebral regions over time. For full legend see Figure 9. Heatmaps of the two genes (D,H) visualizing expression in 14 week old mice.

Figure 12

Graphs showing the mRNA expression for NgR2 (A–C) and NgR3 (E–G) in the 18 cerebral regions over time. For full legend see Figure 9. Heatmaps of the two genes (D,H) visualizing expression in 14 week old mice.

Figure 13

Graphs showing the mRNA expression for Olfactomedin (A–C), LgI1 (E–G) and ADAM22 (I–K) in 18 cerebral regions over time. For full legend see Figure 9. Heatmaps of the three genes (D,H,L) visualizing expression in 14 week old mice.