Biology subjects

Spijker, S.

Publications and source records attributed to Spijker, S..

5 recordsLinked to original sources

bioRxiv2025

A Tiered Approach to Human Synapse Proteomics: Optimized LC-MS/MS Analysis of Whole-Tissue and Synaptosome Preparations from Frozen Post-mortem Brain Samples

Recent advancements in neuroproteomics have enabled detailed analysis of protein expression and function in the human brain. Post-mortem human brain studies have significantly advanced our understanding of the relationship between genetics, cell biology of neurological and psychiatric disorders and their clinical diagnosis. Synaptic dysfunction often has a central role in these disorders. Therefore, we specifically evaluated the sensitivity of liquid chromatography-tandem mass spectrometry (LC-MS/MS) to detect synaptic proteins in whole-tissue lysates versus synaptosome preparations. First, we optimized sample preparation protocols for frozen human gray matter (GM), refining the suspension TRAPping (sTRAP) digestion method to improve protein solubilization and Cysteine reduction and alkylation using thin human tissue sections, and to accomplish low technical variation by minimizing sample handling. We achieved a highly reproducible sample preparation workflow by rigorously applying standardization and randomization across dissection, processing, and LC-MS/MS runs. Second, comparative LC-MS/MS analysis showed that cortical whole-tissue lysates are a practical solution for large-scale studies and broadly detected synaptic proteins focusing on excitatory neurons. However, enrichment by synaptosome isolation offered improved resolution of synapse-specific proteins. Because synapse-proteomics enables insight into spatial regulation--i.e., alterations at the synapse that are not reflected in the soma-we recommend a tiered approach: initial whole-tissue analysis for broad disease-associated changes, followed by targeted synaptosome proteomics to deepen insight into synaptic alterations. This strategy optimally balances throughput, reproducibility, and biological relevance, and enhances the study of brain disorders through proteomics. Moreover, analyzing synaptic proteins first at the tissue level improves insight into overall regulation of synaptic proteins induced by synapse loss or gain.

neuroscience↗

bioRxiv2021

Interfering with contextual fear memories by post-reactivation administration of propranolol in mice: a series of null findings

Post-reactivation amnesia of contextual fear memories by blockade of noradrenergic signaling has been shown to have limited replicability in rodents. This is usually attributed to several boundary conditions that gate the destabilization of memory during its retrieval. How these boundary conditions can be overcome, and what neural mechanisms underlie post-reactivation changes in contextual fear memories remain largely unknown. Here, we report a series of experiments in a contextual fear-conditioning paradigm in mice, that were aimed at solving these issues. We first attempted to obtain a training paradigm that would consistently result in contextual fear memory that could be destabilized upon reactivation, enabling post-retrieval amnesia by administration of propranolol. Unexpectedly, our attempts were unsuccessful to this end. Specifically, over a series of experiments in which we varied different parameters of the fear acquisition procedure, at best small and inconsistent effects were observed. Additionally, we found that propranolol did not alter retrieval-induced neural activity, as measured by the number of c-Fos+ cells in the hippocampal dentate gyrus. To determine whether propranolol was perhaps ineffective in interfering with reactivated contextual fear memories, we also included anisomycin (i.e., a potent and well-known amnesic drug) in several experiments, and measures of synaptic glutamate receptor subunit GluA2 (i.e., a marker of memory destabilization). No post-retrieval amnesia by anisomycin and no altered GluA2 expression by reactivation was observed, suggesting that the memories did not undergo destabilization. The null findings are surprising, given that the training paradigms we implemented were previously shown to result in memories that could be modified upon reactivation. Together, our observations illustrate the elusive nature of reactivation-dependent changes of non-human fear memory.

animal behavior and cognition↗

bioRxiv2019

From stress to depression: Development of extracellular matrix-dependent cognitive impairment following social stress

Stress can predispose to depressive episodes, yet the molecular mechanisms regulating the transition from the initial stress response to a persistent pathological depressive state remain poorly understood. To shed light on this stress-to-depression transition process, we profiled the development of an enduring depressive-like state in rat by assessing affective behavior and hippocampal function during the 2 months following social defeat stress. In addition, we measured remodeling of hippocampal extracellular matrix (ECM) during this period, as we recently identified ECM changes to mediate cognitive impairment during a sustained depressive-like state. We found affective disturbance and cognitive impairment to develop disparately after social stress. While affective deficits emerged gradually, spatial memory impairment was present both early after stress and during the late-emerging chronic depressive-like state. Surprisingly, these phases were separated by a period of normalized hippocampal function. Similarly, the SDPS paradigm induced a biphasic regulation of the hippocampal ECM coinciding with hippocampus-dependent memory deficits. Early after stress, synaptic ECM proteins and the number of perineuronal nets enwrapping parvalbumin-expressing interneurons were decreased. This was followed by a recovery period without ECM dysregulation, before subsequent decreased metalloproteinase activity and ECM build-up, previously shown to impair memory. This suggests that intact hippocampal function requires unaltered ECM levels. Together our data 1) reveal a dichotomy between affective and cognitive impairments similar to that observed in patients, 2) indicate different molecular processes taking place during early stress and the chronic depressive-like state, and 3) support a role of the ECM in mediating long-lasting memory-effects of social stress.

neuroscience↗

bioRxiv2018

Gpr158 deficiency impacts hippocampal CA1 neuronal excitability, dendritic architecture, and affects spatial learning

G-protein-coupled receptor 158 (Gpr158) is highly expressed in striatum, hippocampus and prefrontal cortex. It gained attention as it was implicated in physiological responses to stress and depression. Recently, Gpr158 has been shown to act as a pathway-specific synaptic organizer in the hippocampus, required for proper mossy fiber-CA3 neurocircuitry establishment, structure, and function. Although rodent Gpr158 expression is highest in CA3, considerable expression occurs in CA1 especially after the first postnatal month. Here, we combined hippocampal-dependent behavioral paradigms with subsequent electrophysiological and morphological analyses from the same group of mice to assess the effects of Gpr158 deficiency on CA1 physiology and function. We demonstrate deficits in spatial memory acquisition and retrieval in the Morris water maze paradigm, along with deficits in the acquisition of extinction memory in the passive avoidance test in Gpr158 KO mice. Electrophysiological recordings from CA1 pyramidal neurons revealed normal basal excitatory and inhibitory synaptic transmission, however, Schaffer collateral stimulation yielded dramatically reduced post-synaptic currents. Interestingly, intrinsic excitability of CA1 pyramidals was found increased, potentially acting as a compensatory mechanism to the reductions in Schaffer collateral-mediated drive. Both ex vivo and in vitro, neurons deficient for Gpr158 exhibited robust reductions in dendritic architecture and complexity, i.e., reduced length, surface, bifurcations, and branching. This effect was localized in the apical but not basal compartment of adult CA1 pyramidals, indicative of pathway-specific alterations. A significant positive correlation between spatial memory acquisition and extent of complexity of CA1 pyramidals was found. Taken together, we provide first evidence of significant disruptions in hippocampal CA1 neuronal dendritic architecture and physiology, driven by Gpr158 deficiency. Importantly, the hippocampal neuronal morphology deficits appear to support the impairments in spatial memory acquisition observed in Gpr158 KO mice.

neuroscience↗

bioRxiv2018

Stress vulnerability promotes an alcohol prone phenotype in a preclinical model of sustained depression

Major depression and alcohol-related disorders frequently co-occur. Depression severity weighs on the magnitude and persistence of comorbid alcohol use disorder (AUD), with severe implications for disease prognosis. Here, we investigated whether depression vulnerability drives propensity to AUD at the preclinical level. We used the social defeat-induced persistent stress (SDPS) model of chronic depression in combination with operant alcohol self-administration (SA). Male Wistar rats were subjected to social defeat (5 episodes) and prolonged social isolation (~12 weeks) and subsequently classified as SDPS-prone or SDPS-resilient based on their affective and cognitive performance. Using an operant alcohol SA paradigm, acquisition, motivation, extinction and cue-induced reinstatement of alcohol-seeking were examined in the two subpopulations. SDPS-prone animals showed increased alcohol SA, excessive motivation to acquire alcohol, persistent alcohol-seeking despite alcohol unavailability, extinction resistance and increased cue-induced relapse; the latter could be blocked by the 2 adrenoreceptor agonist guanfacine. In SDPS-resilient rats, prior exposure to social defeat increased alcohol SA without affecting any other measures of alcohol-seeking and -taking. Our data revealed that depression proneness confers vulnerability to alcohol, emulating patterns of alcohol dependence seen in human addicts, and that depression resilience to a large extent protects from the development of AUD-like phenotypes. Furthermore, our data suggest that stress exposure alone, independently of depressive symptoms, alters alcohol intake in the long-term.

animal behavior and cognition↗