Amplified neuroinflammatory responses following an immune challenge occur with normal aging and can elicit or exacerbate neuropathology. The mechanisms mediating this sensitized or "primed" immune ...response in the aged brain are not fully understood. The alarmin high mobility group box 1 (HMGB1) can be released under chronic pathological conditions and initiate inflammatory cascades. This led us to investigate whether HMGB1 regulates age-related priming of the neuroinflammatory response. Here, we show that HMGB1 protein and mRNA were elevated in the hippocampus of unmanipulated aged rats (24-month-old F344XBN rats). Furthermore, aged rats had increased HMGB1 in the CSF, suggesting increased HMGB1 release. We demonstrate that blocking HMGB1 signaling with an intracisterna magna (ICM) injection of the competitive antagonist to HMGB1, Box-A, downregulates basal expression of several inflammatory pathway genes in the hippocampus of aged rats. This indicates that blocking the actions of HMGB1 might reduce age-associated inflammatory priming. To test this hypothesis, we evaluated whether HMGB1 antagonism blocks the protracted neuroinflammatory and sickness response to peripheral Escherichia coli (E. coli) infection in aged rats. ICM pretreatment of aged rats with Box-A 24 h before E. coli infection prevented the extended hippocampal cytokine response and associated cognitive and affective behavioral changes. ICM pretreatment with Box-A also inhibited aging-induced potentiation of the microglial proinflammatory response to lipopolysaccharide ex vivo Together, these results suggest that HMGB1 mediates neuroinflammatory priming in the aged brain. Blocking the actions of HMGB1 appears to "desensitize" aged microglia to an immune challenge, thereby preventing exaggerated behavioral and neuroinflammatory responses following infection.
The world's population is aging, highlighting a need to develop treatments that promote quality of life in aged individuals. Normal aging is associated with precipitous drops in cognition, typically following events that induce peripheral inflammation (e.g., infection, surgery, heart attack). Peripheral immune stimuli cause exaggerated immune responses in the aged brain, which likely underlie these behavioral deficits. Here, we investigated whether the alarmin high mobility group box 1 (HMGB1) mediates age-associated "priming" of the neuroinflammatory response. HMGB1 is elevated in aged rodent brain and CSF. Blocking HMGB1 signaling downregulated expression of inflammatory pathway genes in aged rat brain. Further, HMGB1 antagonism prevented prolonged infection-induced neuroinflammatory and sickness responses in aged rats. Overall, blocking HMGB1 "desensitized" microglia in the aged brain, thereby preventing pathological infection-elicited neuroinflammatory responses.
Highlights • There are temporal differences in sickness behavior in rats. • Changes in sickness behavior are reflected in hippocampal cytokine and microglial responses. • Microglia show dramatic ...differences in immune activation throughout the day, with peak activation during the light phase. • Microglia rhythms are entrained by, but oscillate independent of, glucocorticoids. • These results suggest time-of-day should be considered when planning procedures that induce neuroinflammation.
Abstract Aged animals exhibit diminished circadian rhythms, and both aging and circadian disruption sensitize neuroinflammatory responses. Microglia –the innate immune cell of the CNS – possess ...endogenous timekeeping mechanisms that regulate immune responses. Here, we explored whether aging is associated with disrupted diurnal rhythms in microglia and neuroinflammatory processes. First, hippocampal microglia isolated from young rats (4 mos. F344XBN) rhythmically expressed circadian clock genes, whereas microglia isolated from the hippocampus of aged rats (25 mos.) had aberrant Per1 and Per2 rhythms. Unstimulated microglia from young rats exhibited robust rhythms of TNFα and IL-1β mRNA expression, whereas those from aged rats had flattened and tonically-elevated cytokine expression. Similarly, microglial activation markers were diurnally regulated in the hippocampus of young but not aged rats and diurnal differences in responsiveness to both ex vivo and in vivo inflammatory challenges were abolished in aged rats. Corticosterone is an entraining signal for extra-SCN circadian rhythms. Here, corticosterone stimulation elicited similar Per1 induction in aged and young microglia. Overall, these results indicate that aging dysregulates circadian regulation of neuroinflammatory functions.
Postoperative cognitive dysfunction (POCD) is the collection of cognitive impairments, lasting days to months, experienced by individuals following surgery. Persistent POCD is most commonly ...experienced by older individuals and is associated with a greater vulnerability to developing Alzheimer’s disease, but the underlying mechanisms are not known. It is known that laparotomy (exploratory abdominal surgery) in aged rats produces memory impairments for 4 days. Here we report that postsurgical treatment with morphine extends this deficit to at least 2 months while having no effects in the absence of surgery. Indeed, hippocampal-dependent long-term memory was impaired 2, 4, and 8 weeks postsurgery only in aged, morphine-treated rats. Short-term memory remained intact. Morphine is known to have analgesic effects via μ-opioid receptor activation and neuroinflammatory effects through Toll-like receptor 4 activation. Here we demonstrate that persistent memory deficits were mediated independently of the μ-opioid receptor, suggesting that they were evoked through a neuroinflammatory mechanism and unrelated to pain modulation. In support of this, aged, laparotomized, and morphine-treated rats exhibited increased gene expression of various proinflammatory markers (IL-1β, IL-6, TNFα, NLRP3, HMGB1, TLR2, and TLR4) in the hippocampus at the 2-week time point. Furthermore, central blockade of IL-1β signaling with the specific IL-1 receptor antagonist (IL-1RA), at the time of surgery, completely prevented the memory impairment. Finally, synaptophysin and PSD95 gene expression were significantly dysregulated in the hippocampus of aged, laparotomized, morphine-treated rats, suggesting that impaired synaptic structure and/or function may play a key role in this persistent deficit. This instance of long-term memory impairment following surgery closely mirrors the timeline of persistent POCD in humans and may be useful for future treatment discoveries.
•Surgery and morphine-induced memory deficits lasting 8 weeks in aged, but not young rats.•Morphine-induced POCD in aged rats occurred independently of opioid receptors.•Morphine-induced POCD was associated with increased hippocampal proinflammatory markers.•IL-1RA pretreatment prevented morphine-induced POCD in aged rats.•Morphine-induced POCD was associated with dysregulated hippocampal synaptic markers.
Formation of long-range axons occurs over multiple stages of morphological maturation. However, the intrinsic transcriptional mechanisms that temporally control different stages of axon projection ...development are unknown. Here, we addressed this question by studying the formation of mouse serotonin (5-HT) axons, the exemplar of long-range profusely arborized axon architectures. We report that LIM homeodomain factor 1b (Lmx1b)-deficient 5-HT neurons fail to generate axonal projections to the forebrain and spinal cord. Stage-specific targeting demonstrates that Lmx1b is required at successive stages to control 5-HT axon primary outgrowth, selective routing, and terminal arborization. We show a Lmx1b→Pet1 regulatory cascade is temporally required for 5-HT arborization and upregulation of the 5-HT axon arborization gene, Protocadherin-alphac2, during postnatal development of forebrain 5-HT axons. Our findings identify a temporal regulatory mechanism in which a single continuously expressed transcription factor functions at successive stages to orchestrate the progressive development of long-range axon architectures enabling expansive neuromodulation.
Assembly of transcriptomes encoding unique neuronal identities requires selective accessibility of transcription factors to cis-regulatory sequences in nucleosome-embedded postmitotic chromatin. Yet, ...the mechanisms controlling postmitotic neuronal chromatin accessibility are poorly understood. Here, we show that unique distal enhancers define the
neuron lineage that generates serotonin (5-HT) neurons in mice. Heterogeneous single-cell chromatin landscapes are established early in postmitotic
neurons and reveal the putative regulatory programs driving
neuron subtype identities. Distal enhancer accessibility is highly dynamic as
neurons mature, suggesting the existence of regulatory factors that reorganize postmitotic neuronal chromatin. We find that Pet1 and Lmx1b control chromatin accessibility to select
-lineage-specific enhancers for 5-HT neurotransmission. Additionally, these factors are required to maintain chromatin accessibility during early maturation suggesting that postmitotic neuronal open chromatin is unstable and requires continuous regulatory input. Together, our findings reveal postmitotic transcription factors that reorganize accessible chromatin for neuron specialization.
Vacuolar protein sorting 10 protein (VPS10P) domain receptors are a unique class of intracellular sorting receptors that emerge as major risk factors associated with psychiatric and neurodegenerative ...diseases, including bipolar disorders, autism, schizophrenia, as well as Alzheimer’s disease and frontotemporal dementia. Yet, the lack of suitable experimental models to study receptor functions in the human brain has hampered elucidation of receptor actions in brain disease. Here, we have adapted protocols using human cerebral organoids to the detailed characterization of VPS10P domain receptor expression during neural development and differentiation, including single-cell RNA sequencing. Our studies uncovered spatial and temporal patterns of expression unique to individual receptor species in the human brain. While
SORL1
expression is abundant in stem cells and
SORCS1
peaks in neural progenitors at onset of neurogenesis,
SORT1
and
SORCS2
show increasing expression with maturation of neuronal and non-neuronal cell types, arguing for distinct functions in development
versus
the adult brain. In neurons, subcellular localization also distinguishes between types of receptor species, either mainly localized to the cell soma (
SORL1
and
SORT1
) or also to neuronal projections (
SORCS1
and
SORCS2
), suggesting divergent functions in protein sorting between Golgi and the endo-lysosomal system or along axonal and dendritic tracks. Taken together, our findings provide an important resource on temporal, spatial, and subcellular patterns of VPS10P domain receptor expression in cerebral organoids for further elucidation of receptor (dys) functions causative of behavioral and cognitive defects of the human brain.
Highlights • Timing of stressor exposure influences neuroinflammatory priming. • Microglia inflammatory responses are similarly regulated by time of stress exposure. • The corticosterone response is ...enhanced following stress during the light phase. • Glucocorticoids directly down-regulate genes involved in microglia inhibition.
Abstract Healthy aging individuals are more likely to suffer profound memory impairments following an immune challenge than are younger adults. These challenges produce a brain inflammatory response ...that is exaggerated with age. Sensitized microglia found in the normal aging brain are responsible for this amplified response, which in turn interferes with processes involved in memory formation. Here, we examine factors that may lead aging to sensitize microglia. Aged rats exhibited higher corticosterone levels in the hippocampus, but not in plasma, throughout the daytime (diurnal inactive phase). These elevated hippocampal corticosterone levels were associated with increased hippocampal 11β-hydroxysteroid dehydrogenase type 1 protein expression, the enzyme that catalyzes glucocorticoid formation and greater hippocampal glucocorticoid receptor (GR) activation. Intracisternal administration of mifepristone, a GR antagonist, effectively reduced immune-activated proinflammatory responses, specifically from hippocampal microglia and prevented Escherichia coli –induced memory impairments in aged rats. Voluntary exercise as a therapeutic intervention significantly reduced total hippocampal GR expression. These data strongly suggest that increased GR activation in the aged hippocampus plays a critical role in sensitizing microglia.
Neurons must function for decades of life, but how these non-dividing cells are preserved is poorly understood. Using mouse serotonin (5-HT) neurons as a model, we report an adult-stage ...transcriptional program specialized to ensure the preservation of neuronal connectivity. We uncover a switch in Lmx1b and Pet1 transcription factor function from controlling embryonic axonal growth to sustaining a transcriptomic signature of 5-HT connectivity comprising functionally diverse synaptic and axonal genes. Adult-stage deficiency of Lmx1b and Pet1 causes slowly progressing degeneration of 5-HT synapses and axons, increased susceptibility of 5-HT axons to neurotoxic injury, and abnormal stress responses. Axon degeneration occurs in a die back pattern and is accompanied by accumulation of α-synuclein and amyloid precursor protein in spheroids and mitochondrial fragmentation without cell body loss. Our findings suggest that neuronal connectivity is transcriptionally protected by maintenance of connectivity transcriptomes; progressive decay of such transcriptomes may contribute to age-related diseases of brain circuitry.
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•Lmx1b and Pet1 sustain an adult-stage connectivity transcriptome in 5-HT neurons•Adult-stage loss of Lmx1b and Pet1 causes degeneration of 5-HT synapses and axons•APP and α-syn accumulate in spheroids and mitochondria fragment as fibers degenerate•Connectivity decay is accompanied by physiological and behavioral alterations
Kitt et al. show that adult serotonergic connectivity is protected from expansive degeneration by an Lmx1b/Pet1-regulated transcriptional program. Adult-stage deficiency of these transcription factors causes progressive breakdown of 5-HT synapses and axons accompanied by accumulation of α-synuclein and amyloid precursor protein in spheroids and mitochondrial fragmentation without cell body loss.
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