Browsing by Author "Dudek, Serena M."
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- Hippocampal Subregions Express Distinct Dendritic Transcriptomes that Reveal Differences in Mitochondrial Function in CA2Farris, Shannon; Ward, James M.; Carstens, Kelly E.; Samadi, Mahsa; Wang, Yu; Dudek, Serena M. (CellPress, 2019-10-08)RNA localization is one mechanism neurons use to spatially and temporally regulate gene expression at synapses. Here, we test the hypothesis that cells exhibiting distinct forms of synaptic plasticity will have differences in dendritically localized RNAs. Indeed, we discover that each major subregion of the adult mouse hippocampus expresses a unique complement of dendritic RNAs. Specifically, we describe more than 1,000 differentially expressed dendritic RNAs, suggesting that RNA localization and local translation are regulated in a cell typespecific manner. Furthermore, by focusing Gene Ontology analyses on the plasticity-resistant CA2, we identify an enrichment of mitochondria-associated pathways in CA2 cell bodies and dendrites, and we provide functional evidence that these pathways differentially influence plasticity and mitochondrial respiration in CA2. These data indicate that differences in dendritic transcriptomes may regulate cell type-specific properties important for learning and memory and may influence region-specific differences in disease pathology.
- Histone Hypervariants H2A.Z.1 and H2A.Z.2 Play Independent and Context-Specific Roles in Neuronal Activity-Induced Transcription of Arc/ Arg3.1 and Other Immediate Early GenesDunn, Carissa J.; Sarkar, Pushpita; Bailey, Emma R.; Farris, Shannon; Zhao, Meilan; Ward, James M.; Dudek, Serena M.; Saha, Ramendra N. (Society for Neuroscience, 2017)The histone variant H2A.Z is an essential and conserved regulator of eukaryotic gene transcription. However, the exact role of this histone in the transcriptional process remains perplexing. In vertebrates, H2A.Z has two hypervariants, H2A.Z.1 and H2A.Z.2, that have almost identical sequences except for three amino acid residues. Due to such similarity, functional specificity of these hypervariants in neurobiological processes, if any, remain largely unknown. In this study with dissociated rat cortical neurons, we asked if H2A.Z hypervariants have distinct functions in regulating basal and activity-induced gene transcription. Hypervariant-specific RNAi and microarray analyses revealed that H2A.Z.1 and H2A.Z.2 regulate basal expression of largely nonoverlapping gene sets, including genes that code for several synaptic proteins. In response to neuronal activity, rapid transcription of our model gene Arc is impaired by depletion of H2A.Z.2, but not H2A.Z.1. This impairment is partially rescued by codepletion of the H2A.Z chaperone, ANP32E. In contrast, under a different context (after 48 h of tetrodotoxin, TTX), rapid transcription of Arc is impaired by depletion of either hypervariant. Such context-dependent roles of H2A.Z hypervariants, as revealed by our multiplexed gene expression assays, are also evident with several other immediate early genes, where regulatory roles of these hypervariants vary from gene to gene under different conditions. Together, our data suggest that H2A.Z hypervariants have context-specific roles that complement each other to mediate activity-induced neuronal gene transcription.
- LTD Induction in Adult Visual Cortex: Role of Stimulus Timing and InhibitionPerrett, Stephen P.; Dudek, Serena M.; Eagleman, David M.; Montague, P. Read; Friedlander, Michael J. (Society for Neuroscience, 2001-04-01)One Hertz stimulation of afferents for 15 min with constant interstimulus intervals (regular stimulation) can induce longterm depression (LTD) of synaptic strength in the neocortex. However, it is unknown whether natural patterns of lowfrequency afferent spike activity induce LTD. Although neurons in the neocortex can fire at overall rates as low as 1 Hz, the intervals between spikes are irregular. This irregular spike activity (and thus, presumably, irregular activation of the synapses of that neuron onto postsynaptic targets) can be approximated by stimulation with Poisson-distributed interstimulus intervals (Poisson stimulation). Therefore, if low-frequency presynaptic spike activity in the intact neocortex is sufficient to induce a generalized LTD of synaptic transmission, then Poisson stimulation, which mimics this spike activity, should induce LTD in slices. We tested this hypothesis by comparing changes in the strength of synapses onto layer 2/3 pyramidal cells induced by regular and Poisson stimulation in slices from adult visual cortex. We find that regular stimulation induces LTD of excitatory synaptic transmission as assessed by field potentials and intracellular postsynaptic potentials (PSPs) with inhibition absent. However, Poisson stimulation does not induce a net LTD of excitatory synaptic transmission. When the PSP contained an inhibitory component, neither Poisson nor regular stimulation induced LTD. We propose that the short bursts of synaptic activity that occur during a Poisson train have potentiating effects that offset the induction of LTD that is favored with regular stimulation. Thus, natural (i.e., irregular) low-frequency activity in the adult neocortex in vivo should not consistently induce LTD.
- Mechanisms of mGluR-dependent plasticity in hippocampal area CA2Samadi, Mahsa; Hales, Claire A.; Lustberg, Daniel J.; Farris, Shannon; Ross, Madeleine R.; Zhao, Meilan; Hepler, John R.; Harbin, Nicholas H.; Robinson, Emma S. J.; Banks, Paul J.; Bashir, Zafar I.; Dudek, Serena M. (Wiley, 2023-06)Pyramidal cells in hippocampal area CA2 have synaptic properties that are distinct from the other CA subregions. Notably, this includes a lack of typical long-term potentiation of stratum radiatum synapses. CA2 neurons express high levels of several known and potential regulators of metabotropic glutamate receptor (mGluR)-dependent signaling including Striatal-Enriched Tyrosine Phosphatase (STEP) and several Regulator of G-protein Signaling (RGS) proteins, yet the functions of these proteins in regulating mGluR-dependent synaptic plasticity in CA2 are completely unknown. Thus, the aim of this study was to examine mGluR-dependent synaptic depression and to determine whether STEP and the RGS proteins RGS4 and RGS14 are involved. Using whole cell voltage-clamp recordings from mouse pyramidal cells, we found that mGluR agonist-induced long-term depression (mGluR-LTD) is more pronounced in CA2 compared with that observed in CA1. This mGluR-LTD in CA2 was found to be protein synthesis and STEP dependent, suggesting that CA2 mGluR-LTD shares mechanistic processes with those seen in CA1, but in addition, RGS14, but not RGS4, was essential for mGluR-LTD in CA2. In addition, we found that exogenous application of STEP could rescue mGluR-LTD in RGS14 KO slices. Supporting a role for CA2 synaptic plasticity in social cognition, we found that RGS14 KO mice had impaired social recognition memory as assessed in a social discrimination task. These results highlight possible roles for mGluRs, RGS14, and STEP in CA2-dependent behaviors, perhaps by biasing the dominant form of synaptic plasticity away from LTP and toward LTD in CA2.
- Optimized Method for Robust Transcriptome Profiling of Minute Tissues Using Laser Capture Microdissection and Low-Input RNA-SeqFarris, Shannon; Wang, Yu; Ward, James M.; Dudek, Serena M. (Frontiers, 2017-06-17)Obtaining high quality RNA from complex biological tissues, such as the brain, is needed for establishing high-fidelity cell-type specific transcriptomes. Although combining genetic labeling techniques with laser capture microdissection (LCM) is generally sufficient, concerns over RNA degradation and limited yields call into question results of many sequencing studies. Here we set out to address both of these issues by: (1) developing a fluorescence-assisted LCM protocol that yields high quality RNA from fresh-frozen tissues; and (2) determining a suitable RNA-Seq library generation method for limited amounts of RNA (1–5 ng total RNA). The latter focused on comparing commercially available kits able to produce libraries of sufficient concentration and complexity while limiting PCR amplification biases. We find that high quality RNA (RNA integrity number, RIN, >9) of sufficient concentration can be isolated from lasercaptured material from thinly-sectioned tissues when digestion time and temperature are minimized. Furthermore, we found that library generation approaches that retain ribosomal RNA (rRNA) through cDNA library generation required fewer cycles of PCR, minimizing bias in the resulting libraries. Lastly, end stage depletion of rRNA prior to sequencing enriches for target RNAs, thereby increasing read depth and level of gene detection while decreasing sequencing costs. Here we describe our protocol for generating robust RNA-Seq libraries from laser-captured tissue and demonstrate that with this method, we obtain samples with RNA quality superior to the current standard in the LCM field, and show that low-input RNA-Seq kits that minimize PCR bias produce high fidelity sequencing metrics with less variability compared to current practices.
- Social and novel contexts modify hippocampal CA2 representations of spaceAlexander, Georgia M.; Farris, Shannon; Pirone, Jason R.; Zheng, Chenguang; Colgin, Laura L.; Dudek, Serena M. (Springer Communications, 2016-01-25)The hippocampus supports a cognitive map of space and is critical for encoding declarative memory (who, what, when and where). Recent studies have implicated hippocampal subfield CA2 in social and contextual memory but how it does so remains unknown. Here we find that in adult male rats, presentation of a social stimulus (novel or familiar rat) or a novel object induces global remapping of place fields in CA2 with no effect on neuronal firing rate or immediate early gene expression. This remapping did not occur in CA1, suggesting this effect is specific for CA2. Thus, modification of existing spatial representations might be a potential mechanism by which CA2 encodes social and novel contextual information.