Neuroplasticity across the lifespan : focus on environmental and intrinsic modulators

The plastic brain is unique in its ability to continuously modify its architecture from early development throughout adulthood. Neural plasticity constitutes both the formation of neuronal networks during development and mechanisms that fine-tune synaptic connections for efficient memory consolidation and learning across the lifespan. It becomes more and more clear that although plasticity mechanisms are very similar across life, we can find several stages of plasticity with specific effects on neuronal circuit formation and brain functioning. Prenatal development is mainly genetically driven by programmes orchestrating the transcription of regulatory cues involved in neuronal migration, cell differentiation and morphogenesis. During this stage small signalling molecules like neurotrophins, chemokines and membrane proteins are of major importance during neurogenesis and synaptogenesis. Throughout life, network stabilising mechanisms are crucial for consolidation of long-term memories. Calibration and tuning mechanisms, keeping homeostasis of synaptic levels, ensure stable and efficient integration, processing and storage of information. Dysregulation of plasticity mechanisms can have detrimental effects on higher cognitive functioning, sociability and learning, often manifested in neurodevelopmental disorders.

The gut microbiota-brain axis is an established physiological system referring to the connection of commensal intestinal microbes to the host's brain, which can influence the synaptic strength and neuronal structure via several pathways including microbial metabolites. The bacterial composition in the gut is susceptible to many factors like diet, psychological stress and drugs, which can be reflected in altered levels of released bacterial components and changes in the permeability of the intestinal wall and blood brain barrier (BBB). To investigate the effect of intestinal microbes during the prenatal plasticity phase on functional behaviour in later life, pregnant female mice were overexposed to elevated levels of bacterial cell wall component peptidoglycan (PGN). Female offspring of PGN treated mothers displayed signs of impaired social recognition and motor activity compared to control and male offspring. Functional differences were associated with altered synaptic gene expression between males and females in the prefrontal cortex (PFC). Paper I concludes that exposure to atypically high levels of PGN during late pregnancy might alter prenatal plastic processes, which manifest at juvenile age. Paper II aimed to assess the mechanism behind PGN signalling in immortalised microglia (IMGs) and primary microglia. PGN receptor NOD2 was highly expressed in microglia and upregulated with exposure to increasing PGN levels. NOD2 overactivation stimulated both NF-KB and MAPK downstream signalling, which resulted in increased cytokine and chemokine expression. The first two projects of my thesis show that excessive amounts of maternal PGN can affect prenatal plasticity programmes potentially by changing microglial cytokine and chemokine release, among other possible mechanisms.

Throughout life, experience-dependent neuronal activity is crucial for the formation and strengthening of synapses, which on a structural level, can be measured in spine density and dendritic length dynamics. According to the sleep homeostasis hypothesis (SHY) neuronal connections are strengthened by neuronal activity during the wake period. To restore the net synaptic strength, especially slow wave sleep has been shown to be essential. However, studies investigating the effect of sleep deprivation (SD) on dendritic spine density and length in mice have shown contradictory results. By conducting a meta-analysis of the current literature, Paper III aimed to resolve conflicting findings across studies on the effect of SD on dendritic architecture. We would expect that SD leads to an increase of synaptic numbers per spine length and longer dendrites, since the lack of sleeps means a lack of downregulation in net synaptic strength proposed by the SHY. Combining studies in chronically (> 24 h) sleep deprived animals, dendritic length and spine density was significantly decreased. These results need to be interpreted with caution, since effects were driven mainly by the subgroup using a SD protocol with aversive stimuli with a substantial heterogeneity across studies. To draw a conclusion more studies have to be performed using standardised methods for SD and dendrite analysis. To reduce variability across studies a methodological checklist was established addressing core issues identified during the quality assessment.

During adulthood neuronal networks are typically stable and only susceptible to structural changes during high frequency stimulation. Strong neuronal activation is subsequently leading to a sustained upregulation of synaptic strength, a term known as long-term potentiation (LTP). To open the window of plasticity, plasticity inhibitory systems like the Nogo system need to be downregulated. Paper IV investigates how plasticity stabilisers like the Nogo system are regulated by using pharmacological agents to induce LTP and long-term depression (LTD) in hippocampal primary cultures. Dendritic and cytosolic NgR1 protein was downregulated by LTP after 60 min, which correlated with time of immediate early gene (IEG) induction. LTD upregulated NgR1 transiently (30-60 min). To further investigate specific pathways and genes that target NgR1 regulation, in Paper V, we performed a CRISPR activation screen in NgRKOGFP reporter cells. NgRKOGFP cells, expressing the reporter protein GFP in the NgR1 locus, were transduced with a whole genome sgRNA library to detect genes that up regulate (GFPhigh population) or down regulate (GFPlow population) NgR1 expression. Several pathways associated with activity-dependent signalling were enriched in the GFPlow population using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases for gene set enrichment analysis (GSEA). To dig deeper into LTP-dependent genes involved in NgR1 downregulation, we correlated our GFPlow to a publicly available RiboTag dataset of upregulated genes following cLTP induction in hippocampal slices. Genes upregulated after LTP induction were positively correlated with downregulation of NgR1 expression. Neuron-specific IEG NPAS4 was identified to play a major role in LTP-dependent control of NgR1.

This thesis focuses on different mechanisms engaged in prenatal and postnatal plasticity regulation. By going deeper into environmental factors like the gut microbiota and endogenous systems like sleep and the Nogo system, new potential molecular mechanisms were discovered and existing pathways broadened.

List of scientific papers

I. Martínez Sánchez I, Spielbauer J, Diaz Heijtz R. Maternal peptidoglycan overexposure during late pregnancy alters neurodevelopment and behavior in juvenile offspring. Brain Behav Immun. 2025;127:96-102.
https://doi.org/10.1016/j.bbi.2025.03.014

II. Spielbauer J, Glotfelty EJ, Sarlus H, Harris RA, Diaz Heijtz R, Karlsson TE. Bacterial peptidoglycan signalling in microglia: Activation by MDP via the NF-KB/MAPK pathway. Brain Behav Immun. 2024;121:43-55.
https://doi.org/10.1016/j.bbi.2024.06.027

III. Brodin ATS, Liesecke F, Spielbauer J, Karlsson TE. Sleep deprivation and dendritic architecture: a systematic review and meta-analysis. [Manuscript]

IV. Brodin ATS, Spielbauer J, Wellfelt K, Olson L, Karlsson TE. Localisation and regulation of Nogo-66 receptor 1 during chemical long-term potentiation and depression. [Manuscript]

V. Spielbauer J, Pfeiffer S, Schick JA, Karlsson TE. Genome-wide CRISPR activation screen identifies key regulators of NgR1. [Manuscript]