Ichijo, R;Maki, K;Kabata, M;Murata, T;Nagasaka, A;Ishihara, S;Haga, H;Honda, T;Adachi, T;Yamamoto, T;Toyoshima, F;
| DOI: 10.1038/s43587-022-00244-6
Stem cell loss causes tissue deterioration associated with aging. The accumulation of genomic and oxidative stress-induced DNA damage is an intrinsic cue for stem cell loss1 [/articles/s43587-022-00244-6#ref-CR1],2 [/articles/s43587-022-00244-6#ref-CR2]; however, whether there is an external microenvironmental cue that triggers stem cell loss remains unclear. Here we report that the involution of skin vasculature causes dermal stiffening that augments the differentiation and hemidesmosome fragility of interfollicular epidermal stem cells (IFESCs) in aged mouse skin. Aging-related IFESC dysregulation occurs in plantar and tail skin, and is correlated with prolonged calcium influx, which is contributed by the mechanoresponsive ion channel Piezo1 (ref. 3 [/articles/s43587-022-00244-6#ref-CR3]). Epidermal deletion of Piezo1 ameliorated IFESC dysregulation in aged skin, whereas Piezo1 activation augmented IFESC differentiation and hemidesmosome fragility in young mice. The dermis stiffened with age, which was accompanied by dermal vasculature atrophy. Conversely, induction of the dermal vasculature softened the dermis and ameliorated IFESC dysregulation in aged skin. Single-cell RNA sequencing of dermal fibroblasts identified an aging-associated anti-angiogenetic secretory molecule, pentraxin 3 (ref. 4 [/articles/s43587-022-00244-6#ref-CR4]), which caused dermal sclerotization and IFESC dysregulation in aged skin. Our findings show that the vasculature softens the microenvironment for stem cell maintenance and provide a potential mechanobiology-based therapeutic strategy against skin disorders in aging.
Functional roles for Piezo1 and Piezo2 in urothelial mechanotransduction and lower urinary tract interoception
Dalghi, MG;Ruiz, WG;Clayton, DR;Montalbetti, N;Daugherty, SL;Beckel, JM;Carattino, MD;Apodaca, G;
PMID: 34464353 | DOI: 10.1172/jci.insight.152984
The mechanisms that link visceral mechanosensation to the perception of internal organ status (i.e., interoception) remain elusive. In response to bladder filling, the urothelium releases ATP, which is hypothesized to stimulate voiding function by communicating the degree of bladder fullness to subjacent tissues including afferent nerve fibers. To determine if PIEZO channels function as mechanosensors in these events, we generated conditional urothelial Piezo1-, Piezo2-, and dual Piezo1/2-knockout (KO) mice. While functional PIEZO1 channels were expressed in all urothelial cell layers, Piezo1-KO mice had a limited phenotype. Piezo2 expression was limited to a small subset of superficial umbrella cells, yet male Piezo2-KO mice exhibited incontinence (i.e., leakage) when their voiding behavior was monitored during their active dark phase. Dual Piezo1/2-KO mice had the most significant phenotype, characterized by decreased urothelial responses to mechanical stimulation, diminished ATP release, bladder hypoactivity in anesthetized Piezo1/2-KO females, but not male ones, and urinary incontinence in both male and female Piezo1/2-KO mice during their dark phase, but not inactive light one. Our studies reveal that the urothelium functions in a sex and circadian manner to link urothelial PIEZO1/2 channel-driven mechanotransduction to normal voiding function and behavior, and in the absence of these signals, bladder dysfunction ensues.
Al-Hasani R, McCall JG, Shin G, Gomez AM, Schmitz GP, Bernardi JM, Pyo CO, Park SI, Marcinkiewcz CM, Crowley NA, Krashes MJ, Lowell BB, Kash TL, Rogers JA, Bruchas MR.
PMID: 26335648 | DOI: 10.1016/j.neuron.2015.08.019
The nucleus accumbens (NAc) and the dynorphinergic system are widely implicated in motivated behaviors. Prior studies have shown that activation of the dynorphin-kappa opioid receptor (KOR) system leads to aversive, dysphoria-like behavior. However, the endogenous sources of dynorphin in these circuits remain unknown. We investigated whether dynorphinergic neuronal firing in the NAc is sufficient to induce aversive behaviors. We found that photostimulation of dynorphinergic cells in the ventral NAc shell elicits robust conditioned and real-time aversive behavior via KOR activation, and in contrast, photostimulation of dorsal NAc shell dynorphin cells induced a KOR-mediated place preference and was positively reinforcing. These results show previously unknown discrete subregions of dynorphin-containing cells in the NAc shell that selectively drive opposing behaviors. Understanding the discrete regional specificity by which NAc dynorphinerigic cells regulate preference and aversion provides insight into motivated behaviors that are dysregulated in stress, reward, and psychiatric disease.
Woo SH, Lukacs V, de Nooij JC, Zaytseva D, Criddle CR, Francisco A, Jessell TM, Wilkinson KA, Patapoutian A.
PMID: 26551544 | DOI: 10.1038/nn.4162.
Proprioception, the perception of body and limb position, is mediated by proprioceptors, specialized mechanosensory neurons that convey information about the stretch and tension experienced by muscles, tendons, skin and joints. In mammals, the molecular identity of the stretch-sensitive channel that mediates proprioception is unknown. We found that the mechanically activated nonselective cation channel Piezo2 was expressed in sensory endings of proprioceptors innervating muscle spindles and Golgi tendon organs in mice. Two independent mouse lines that lack Piezo2 in proprioceptive neurons showed severely uncoordinated body movements and abnormal limb positions. Moreover, the mechanosensitivity of parvalbumin-expressing neurons that predominantly mark proprioceptors was dependent on Piezo2 expression in vitro, and the stretch-induced firing of proprioceptors in muscle-nerve recordings was markedly reduced in Piezo2-deficient mice. Together, our results indicate that Piezo2 is the major mechanotransducer of mammalian proprioceptors.
von Schimmelmann M, Feinberg PA, Sullivan JM, Ku SM, Badimon A, Duff MK, Wang Z, Lachmann A, Dewell S, Ma'ayan A, Han MH, Tarakhovsky A, Schaefer A.
PMID: 27526204 | DOI: 10.1038/nn.4360
Normal brain function depends on the interaction between highly specialized neurons that operate within anatomically and functionally distinct brain regions. Neuronal specification is driven by transcriptional programs that are established during early neuronal development and remain in place in the adult brain. The fidelity of neuronal specification depends on the robustness of the transcriptional program that supports the neuron type-specific gene expression patterns. Here we show that polycomb repressive complex 2 (PRC2), which supports neuron specification during differentiation, contributes to the suppression of a transcriptional program that is detrimental to adult neuron function and survival. We show that PRC2 deficiency in striatal neurons leads to the de-repression of selected, predominantly bivalent PRC2 target genes that are dominated by self-regulating transcription factors normally suppressed in these neurons. The transcriptional changes in PRC2-deficient neurons lead to progressive and fatal neurodegeneration in mice. Our results point to a key role of PRC2 in protecting neurons against degeneration.
Godino, A;Salery, M;Durand-de Cuttoli, R;Estill, MS;Holt, LM;Futamura, R;Browne, CJ;Mews, P;Hamilton, PJ;Neve, RL;Shen, L;Russo, SJ;Nestler, EJ;
PMID: 36889314 | DOI: 10.1016/j.neuron.2023.02.013
The complex nature of the transcriptional networks underlying addictive behaviors suggests intricate cooperation between diverse gene regulation mechanisms that go beyond canonical-activity-dependent pathways. Here, we implicate in this process a nuclear receptor transcription factor, retinoid X receptor alpha (RXRα), which we initially identified bioinformatically as associated with addiction-like behaviors. In the nucleus accumbens (NAc) of male and female mice, we show that although its own expression remains unaltered after cocaine exposure, RXRα controls plasticity- and addiction-relevant transcriptional programs in both dopamine receptor D1- and D2-expressing medium spiny neurons, which in turn modulate intrinsic excitability and synaptic activity of these NAc cell types. Behaviorally, bidirectional viral and pharmacological manipulation of RXRα regulates drug reward sensitivity in both non-operant and operant paradigms. Together, this study demonstrates a key role for NAc RXRα in promoting drug addiction and paves the way for future studies of rexinoid signaling in psychiatric disease states.
Matsuo, J;Mon, N;Douchi, D;Yamamura, A;Kulkarni, M;Heng, D;Chen, S;Nuttonmanit, N;Li, Y;Yang, H;Lee, M;Tam, W;Osato, M;Chuang, L;Ito, Y;
| DOI: 10.1093/stmcls/sxab009
Mammary gland homeostasis is maintained by adult tissue stem-progenitor cells residing within the luminal and basal epithelia. Dysregulation of mammary stem cells is a key mechanism for cancer development. However, stem cell characterization is challenging because reporter models using cell-specific promoters do not fully recapitulate the mammary stem cell populations. We previously found that a 270-basepair Runx1 enhancer element, named eR1, marked stem cells in the blood and stomach. Here, we identified eR1 activity in a rare subpopulation of the ERα-negative luminal epithelium in mouse mammary glands. Lineage-tracing using an eR1-CreERT2 mouse model revealed that eR1+ luminal cells generated the entire luminal lineage and milk-secreting alveoli - eR1 therefore specifically marks lineage-restricted luminal stem cells. eR1-targeted-conditional knockout of Runx1 led to the expansion of luminal epithelial cells, accompanied by elevated ERα expression. Our findings demonstrate a definitive role for Runx1 in the regulation of the eR1-positive luminal stem cell proliferation during mammary homeostasis. Our findings identify a mechanistic link for Runx1 in stem cell proliferation and its dysregulation in breast cancer. Runx1 inactivation is therefore likely to be an early hit in the cell-of-origin of ERα+ luminal type breast cancer.
Hennes A, Held K, Boretto M, De Clercq K, Van den Eynde C, Vanhie A, Van Ranst N, Benoit M, Luyten C, Peeraer K, Tomassetti C, Meuleman C, Voets T, Vankelecom H, Vriens J.
PMID: 30741991 | DOI: 10.1038/s41598-018-38376-8
Successful pregnancy requires the establishment of a complex dialogue between the implanting embryo and the endometrium. Knowledge regarding molecular candidates involved in this early communication process is inadequate due to limited access to primary human endometrial epithelial cells (EEC). Since pseudo-pregnancy in rodents can be induced by mechanical scratching of an appropriately primed uterus, this study aimed to investigate the expression of mechanosensitive ion channels in EEC. Poking of EEC provoked a robust calcium influx and induced an increase in current densities, which could be blocked by an inhibitor of mechanosensitive ion channels. Interestingly, RNA expression studies showed high expression of PIEZO1 in EEC of mouse and human. Additional analysis provided further evidence for the functional expression of PIEZO1 since stimulation with Yoda1, a chemical agonist of PIEZO1, induced increases in intracellular calcium concentrations and current densities in EEC. Moreover, the ion channel profile of human endometrial organoids (EMO) was validated as a representative model for endometrial epithelial cells. Mechanical and chemical stimulation of EMO induced strong calcium responses supporting the hypothesis of mechanosensitive ion channel expression in endometrial epithelial cells. In conclusion, EEC and EMO functionally express the mechanosensitive PIEZO1 channel that could act as a potential target for the development of novel treatments to further improve successful implantation processes.
Kim, JS;Williams, KC;Kirkland, RA;Schade, R;Freeman, KG;Cawthon, CR;Rautmann, AW;Smith, JM;Edwards, GL;Glenn, TC;Holmes, PV;de Lartigue, G;de La Serre, CB;
PMID: 37380023 | DOI: 10.1016/j.molmet.2023.101764
Obesity is associated with deficits in reward which have been linked to compensatory overeating. The vagus nerve is a direct neural pathway that conveys post-ingestive feedback from the gut to the brain, including the reward regions, and vagal activation causes stereotypical reward behaviors. Chronic high fat (HF) feeding alters vagal signaling potentially dampening food-associated reward. Microbiota composition changes rapidly with HF feeding, and a HF-type microbiota is sufficient to alter vagal structure and function. However, whether microbiota-driven alterations in vagal signaling affect host appetitive feeding behavior is unknown. Here, we investigate if microbiota composition modulates reward signaling and assess the role of the vagus in mediating microbiota to brain communication. Male germ-free Fisher rats were colonized with gastrointestinal contents from chow (low fat (LF) ConvLF) or HF (ConvHF) fed rats. Following colonization, ConvHF rats consumed significantly more food than ConvLF animals. ConvHF rats displayed lower feeding-induced extracellular DOPAC levels (a metabolite of dopamine) in the Nucleus Accumbens (NAc) as well as reduced motivation for HF foods compared to ConvLF rats. Dopamine receptor 2 (DDR2) expression levels in the NAc were also significantly lower in ConvHF animals. Similar deficits were observed in conventionally raised HF fed rats, showing that diet-driven alteration in reward can be initiated via microbiota. Selective gut to brain deafferentation restored DOPAC levels, DRD2 expression, and motivational drive in ConvHF rats. We concluded from these data that a HF-type microbiota is sufficient to alter appetitive feeding behavior and that bacteria to reward communication is mediated by the vagus nerve.
Reiner, BC;Zhang, Y;Stein, LM;Perea, ED;Arauco-Shapiro, G;Ben Nathan, J;Ragnini, K;Hayes, MR;Ferraro, TN;Berrettini, WH;Schmidt, HD;Crist, RC;
PMID: 36075888 | DOI: 10.1038/s41398-022-02135-1
Opioid exposure is known to cause transcriptomic changes in the nucleus accumbens (NAc). However, no studies to date have investigated cell type-specific transcriptomic changes associated with volitional opioid taking. Here, we use single nucleus RNA sequencing (snRNAseq) to comprehensively characterize cell type-specific alterations of the NAc transcriptome in rats self-administering morphine. One cohort of male Brown Norway rats was injected with acute morphine (10 mg/kg, i.p.) or saline. A second cohort of rats was allowed to self-administer intravenous morphine (1.0 mg/kg/infusion) for 10 consecutive days. Each morphine-experienced rat was paired with a yoked saline control rat. snRNAseq libraries were generated from NAc punches and used to identify cell type-specific gene expression changes associated with volitional morphine taking. We identified 1106 differentially expressed genes (DEGs) in the acute morphine group, compared to 2453 DEGs in the morphine self-administration group, across 27 distinct cell clusters. Importantly, we identified 1329 DEGs that were specific to morphine self-administration. DEGs were identified in novel clusters of astrocytes, oligodendrocytes, and D1R- and D2R-expressing medium spiny neurons in the NAc. Cell type-specific DEGs included Rgs9, Celf5, Oprm1, and Pde10a. Upregulation of Rgs9 and Celf5 in D2R-expressing neurons was validated by RNAscope. Approximately 85% of all oligodendrocyte DEGs, nearly all of which were associated with morphine taking, were identified in two subtypes. Bioinformatic analyses identified cell type-specific upstream regulatory mechanisms of the observed transcriptome alterations and downstream signaling pathways, including both novel and previously identified molecular pathways. These findings show that volitional morphine taking is associated with distinct cell type-specific transcriptomic changes in the rat NAc and highlight specific striatal cell populations and novel molecular substrates that could be targeted to reduce compulsive opioid taking.
Caprioli D, Venniro M, Zhang M, Bossert JM, Warren BL, Hope BT, Shaham Y.
PMID: 27980115 | DOI: 10.1523/JNEUROSCI.3091-16.2016
We recently developed a rat model of incubation of methamphetamine craving after choice-based voluntary abstinence. Here, we studied the role of dorsolateral and dorsomedial striatum (DLS, DMS) in this incubation.We trained rats to self-administer palatable food pellets (6 days, 6-h/d) and methamphetamine (12 days, 6-h/d). We then assessed relapse to methamphetamine seeking under extinction conditions after 1 and 21 abstinence days. Between tests, the rats underwent voluntary abstinence (using a discrete choice procedure between methamphetamine and food; 20 trials/day) for 19 days. We used in situ hybridization to measure co-labeling of the activity marker Fos with Drd1 and Drd2 in DMS and DLS after the tests. Based on the in situ hybridization co-labeling results, we tested the causal role of DMS D1- and D2-family receptors, and DMS neuronal ensembles in 'incubated' methamphetamine seeking, using selective dopamine receptor antagonists (SCH39166 or raclopride) and the Daun02 chemogenetic inactivation procedure, respectively.Methamphetamine seeking was higher after 21 days of voluntary abstinence than after 1 day (incubation of methamphetamine craving). The 'incubated' response was associated with increased Fos expression in DMS but not DLS; Fos was co-labeled with both Drd1 and Drd2 DMS injections of SCH39166 or raclopride selectively decreased methamphetamine seeking after 21 abstinence days. In Fos-lacZ transgenic rats, selective inactivation of relapse test-activated Fos neurons in DMS on abstinence day 18 decreased incubated methamphetamine seeking on day 21.Results demonstrate a role of DMS dopamine D1 and D2-receptors in incubation of methamphetamine craving after voluntary abstinence and that DMS neuronal ensembles mediate this incubation.
SIGNIFICANCE STATEMENT:
In human addicts, abstinence is often self-imposed and relapse can be triggered by exposure to drug-associated cues that induce drug craving. We recently developed a rat model of incubation of methamphetamine craving after choice-based voluntary abstinence. Here, we used classical pharmacology, in situ hybridization, immunohistochemistry, and the Daun02 inactivation procedure to demonstrate a critical role of dorsomedial striatum neuronal ensembles in this new form of incubation of drug craving.
bioRxiv : the preprint server for biology
Zhang, W;Zhao, J;Deng, L;Ishimwe, N;Pauli, J;Wu, W;Shan, S;Kempf, W;Ballantyne, MD;Kim, D;Lyu, Q;Bennett, M;Rodor, J;Turner, AW;Lu, YW;Gao, P;Choi, M;Warthi, G;Kim, HW;Barroso, MM;Bryant, WB;Miller, CL;Weintraub, NL;Maegdefessel, L;Miano, JM;Baker, AH;Long, X;
PMID: 36711681 | DOI: 10.1101/2023.01.07.522948
Activation of vascular smooth muscle cells (VSMCs) inflammation is vital to initiate vascular disease. However, the role of human-specific long noncoding RNAs (lncRNAs) in VSMC inflammation is poorly understood.Bulk RNA-seq in differentiated human VSMCs revealed a novel human-specific lncRNA called IN flammatory M K L1 I nteracting L ong N oncoding RNA ( INKILN ). INKILN expression was assessed in multiple in vitro and ex vivo models of VSMC phenotypic modulation and human atherosclerosis and abdominal aortic aneurysm (AAA) samples. The transcriptional regulation of INKILN was determined through luciferase reporter system and chromatin immunoprecipitation assay. Both loss- and gain-of-function approaches and multiple RNA-protein and protein-protein interaction assays were utilized to uncover the role of INKILN in VSMC proinflammatory gene program and underlying mechanisms. Bacterial Artificial Chromosome (BAC) transgenic (Tg) mice were utilized to study INKLIN expression and function in ligation injury-induced neointimal formation.INKILN expression is downregulated in contractile VSMCs and induced by human atherosclerosis and abdominal aortic aneurysm. INKILN is transcriptionally activated by the p65 pathway, partially through a predicted NF-κB site within its proximal promoter. INKILN activates the proinflammatory gene expression in cultured human VSMCs and ex vivo cultured vessels. Mechanistically, INKILN physically interacts with and stabilizes MKL1, a key activator of VSMC inflammation through the p65/NF-κB pathway. INKILN depletion blocks ILIβ-induced nuclear localization of both p65 and MKL1. Knockdown of INKILN abolishes the physical interaction between p65 and MKL1, and the luciferase activity of an NF-κB reporter. Further, INKILN knockdown enhances MKL1 ubiquitination, likely through the reduced physical interaction with the deubiquitinating enzyme, USP10. INKILN is induced in injured carotid arteries and exacerbates ligation injury-induced neointimal formation in BAC Tg mice.These findings elucidate an important pathway of VSMC inflammation involving an INKILN /MKL1/USP10 regulatory axis. Human BAC Tg mice offer a novel and physiologically relevant approach for investigating human-specific lncRNAs under vascular disease conditions.
