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Frontiers in Neuroscience 2022Importin α3 (Gene: , the ortholog of human Importin α4) is a member of the importin α family and participates in nucleocytoplasmic transport by forming trimeric...
Importin α3 (Gene: , the ortholog of human Importin α4) is a member of the importin α family and participates in nucleocytoplasmic transport by forming trimeric complexes between cargo proteins and importin β1. Evidence from human studies has indicated that single nucleotide polymorphisms (SNP) in the gene are associated with the occurrence of several psychiatric disorders accompanied by abnormal reward-related behavior, including schizophrenia, major depression, and substance addiction. However, the precise roles of importin α3 in controlling reward processing and motivation are still unclear. In this study, we evaluated the behavioral effects of knockout (KO) in mice on performance in touchscreen operant chamber-based tasks evaluating simple (fixed-ratio) and effortful (progressive-ratio) reward-seeking behaviors. While KO mice showed no significant differences in operant reward learning on a fixed-ratio schedule, they demonstrated significantly increased motivation (increased break point) to instrumentally respond for sucrose on a progressive-ratio schedule. We additionally measured the number of c-Fos-positive cells, a marker of neural activity, in 20 regions of the brain and identified a network of brain regions based on their interregional correlation coefficients. Network and graph-theoretic analyses suggested that deficiency enhanced overall interregional functional connectivity. These findings suggest the importance of in motivational control and indicate that KO mice may be an attractive line for modeling motivational abnormalities associated with several psychiatric disorders.
PubMed: 35844217
DOI: 10.3389/fnins.2022.905991 -
Journal of Pharmacological Sciences Jul 2022Osteoporosis is caused by an imbalance in bone remodeling due to abnormal osteoclast (OC) formation and activation. Hypoxia at the site of inflammation promotes OC...
Osteoporosis is caused by an imbalance in bone remodeling due to abnormal osteoclast (OC) formation and activation. Hypoxia at the site of inflammation promotes OC formation and activation in various species, including humans. We previously reported that insulin-like growth factor 2 (IGF2) plays an important role in osteoclastogenesis under hypoxia. In our present study, we focused on the mechanism of osteoclastogenesis in regard to IGF2 signaling under hypoxia. We confirmed that the addition of IGF2 promoted osteoclastogenesis under normoxic conditions. Conversely, IGF2-neutralizing antibodies inhibited osteoclastogenesis under both normoxic and hypoxic conditions. IGF2 addition increased levels of phosphorylated Akt (Thr308 and Ser473) and NF-κB (Ser536), indicating activation of the Akt-NF-κB pathway. IGF2 also increased the expression of inducible nitric oxide synthase, which promotes osteoclastogenesis via nitric oxide production. Expression levels of genes encoding inflammatory cytokines, such as tumor necrosis factor-α, interleukin (IL)-1β, and IL-6, were upregulated, indicating that IGF2 promotes osteoclastogenesis by increasing the expression of inflammatory cytokines via activation of the Akt-NF-κB pathway. These results suggest that IGF2 is a promising therapeutic target for osteoporosis and rheumatoid arthritis.
Topics: Cytokines; Humans; Hypoxia; Insulin-Like Growth Factor II; NF-kappa B; Osteogenesis; Osteoporosis; Proto-Oncogene Proteins c-akt
PubMed: 35641033
DOI: 10.1016/j.jphs.2022.03.007 -
Journal of Neuroinflammation May 2020Hypoxic-ischemic encephalopathy (HIE) has a high morbidity rate and involves severe neurologic deficits, including cerebral palsy. Therapeutic hypothermia (TH) has been...
BACKGROUND
Hypoxic-ischemic encephalopathy (HIE) has a high morbidity rate and involves severe neurologic deficits, including cerebral palsy. Therapeutic hypothermia (TH) has been shown to decrease the mortality rate and provide neuroprotection in infants with HIE. However, death and disability rates in HIE infants treated with TH remain high. Although the cellular mechanism of the neuroprotective effect of TH remains unclear, astrocytic erythropoietin (EPO) is known to be a key mediator of neuroprotection under hypoxic conditions. In the present study, we investigated the hypothermia effect on EPO expression in astrocytes and determined whether hypothermia attenuates neuronal damage via EPO signaling.
METHODS
Astrocytes derived from rat cerebral cortex were cultured under oxygen/glucose deprivation (OGD). The expression of EPO and hypoxia-inducible factor (HIF), a transcription factor of EPO, was assessed. After OGD, astrocytes were cultured under normothermic (37 °C) or hypothermic (33.5 °C) conditions, and then EPO and HIF expression was assessed. After OGD, rat cortical neurons were cultured in astrocyte-conditioned medium (ACM) derived from the hypothermic group, and neuronal apoptosis was evaluated.
RESULTS
OGD induced EPO mRNA and protein expression, although at lower levels than hypoxia alone. HIF-1α and HIF-2α protein expression increased under hypoxia alone and OGD, although OGD increased HIF-2α protein expression less than hypoxia alone. EPO gene and protein expression after OGD was significantly higher under hypothermia. Moreover, expression of HIF-1α and HIF-2α protein was enhanced under hypothermia. In the presence of ACM derived from hypothermic astrocytes following OGD, the number of cleaved caspase 3 and TdT-mediated dUTP nick-end labeling-positive apoptotic neurons was lower than in the presence of ACM from normothermic astrocytes following OGD. Blockade of EPO signaling using anti-EPO neutralization antibody attenuated the anti-apoptotic effect of ACM derived from hypothermic astrocytes following OGD.
CONCLUSIONS
Hypothermia after OGD stabilized HIF-EPO signaling in astrocytes, and upregulated EPO expression could suppress neuronal apoptosis. Investigating the neuroprotective effect of EPO from astrocytes under hypothermic conditions may contribute to the development of novel neuroprotection-based therapies for HIE.
Topics: Animals; Astrocytes; Erythropoietin; Hypothermia, Induced; Hypoxia-Ischemia, Brain; Neurons; Neuroprotection; Rats; Rats, Wistar
PubMed: 32359362
DOI: 10.1186/s12974-020-01831-3 -
Journal of Pharmacological Sciences Jan 2023Bone remodeling mediated by bone-forming osteoblasts (OBs) and bone-resorbing osteoclasts (OCs) maintains bone structure and function. Excessive OC activation leads to...
Bone remodeling mediated by bone-forming osteoblasts (OBs) and bone-resorbing osteoclasts (OCs) maintains bone structure and function. Excessive OC activation leads to bone-destroying diseases such as osteoporosis and bone erosion of rheumatoid arthritis (RA). Differentiation of OCs from bone marrow cells (BMCs) is regulated by the bone microenvironment. The proinflammatory cytokine interleukin (IL)-1β reportedly enhances osteoclastogenesis and plays important roles in RA-associated bone loss. The present study investigated the effect of IL-1β on OC formation via microenvironmental cells. Treating mouse BMCs with IL-1β in the presence of receptor activator of NF-κB ligand and macrophage colony-stimulating factor increased the number of OCs. Real-time RT-PCR revealed increased expression of the IL-1β, IL-1RI, and IL-1RII genes in non-OCs compared with OCs. Removing CD45 cells which cannot differentiate into OCs, from mouse BMCs reduced the IL-1β-mediated enhancement of osteoclastogenesis. IL-1β treatment upregulated the expression of inducible nitric oxide synthase, insulin-like growth factor 2 (IGF2), and the chemokines stromal cell derived factor 1, C-X3-C motif ligand 1 (CX3CL1), and CXCL7 in non-OCs. Neutralizing antibodies against these chemokines and IGF2 suppressed osteoclastogenesis in the presence of IL-1β. These results suggest that IL-1β enhances osteoclastogenesis by upregulating IGF2 and chemokine expression in non-OCs.
Topics: Mice; Animals; Osteogenesis; Ligands; Cells, Cultured; Osteoclasts; Osteoblasts; Cell Differentiation; RANK Ligand
PubMed: 36522118
DOI: 10.1016/j.jphs.2022.10.007 -
Translational Oncology Feb 2021M2-like tumor-associated macrophages (M2-TAMs) in cancer tissues are intimately involved in cancer immunosuppression in addition to growth, invasion, angiogenesis, and...
M2-like tumor-associated macrophages (M2-TAMs) in cancer tissues are intimately involved in cancer immunosuppression in addition to growth, invasion, angiogenesis, and metastasis. Hence, considerable attention has been focused on cancer immunotherapies targeting M2-TAMs. However, systemic therapies inhibit TAMs as well as other macrophages important for normal immune responses throughout the body. To stimulate tumor immunity with fewer side effects, we targeted M2-TAMs using photodynamic therapy (PDT), which damages cells via a nontoxic photosensitizer with harmless laser irradiation. We synthesized a light-sensitive compound, mannose-conjugated chlorin e6 (M-chlorin e6), which targets mannose receptors highly expressed on M2-TAMs. M-chlorin e6 accumulated more in tumor tissue than normal skin tissue of syngeneic model mice and was more rapidly excreted than the second-generation photosensitizer talaporfin sodium. Furthermore, M-chlorin e6 PDT significantly reduced the volume and weight of tumor tissue. Flow cytometric analysis revealed that M-chlorin e6 PDT decreased the proportion of M2-TAMs and increased that of anti-tumor macrophages, M1-like TAMs. M-chlorin e6 PDT also directly damaged and killed cancer cells in vitro. Our data indicate that M-chlorin e6 is a promising new therapeutic agent for cancer PDT.
PubMed: 33401079
DOI: 10.1016/j.tranon.2020.101005 -
The American Journal of Pathology Dec 2021Bone homeostasis depends on the balance between bone resorption by osteoclasts (OCs) and bone formation by osteoblasts. Bone resorption can become excessive under...
Bone homeostasis depends on the balance between bone resorption by osteoclasts (OCs) and bone formation by osteoblasts. Bone resorption can become excessive under various pathologic conditions, including rheumatoid arthritis. Previous studies have shown that OC formation is promoted under hypoxia. However, the precise mechanisms behind OC formation under hypoxia have not been elucidated. The present study investigated the role of inducible nitric oxide synthase (iNOS) in OC differentiation under hypoxia. Primary bone marrow cells obtained from mice were stimulated with receptor activator of NF-κB ligand and macrophage colony-stimulating factor to induce OC differentiation. The number of OCs increased in culture under hypoxia (oxygen concentration, 5%) compared with that under normoxia (oxygen concentration, 20%). iNOS gene and protein expression increased in culture under hypoxia. Addition of an iNOS inhibitor under hypoxic conditions suppressed osteoclastogenesis. Addition of a nitric oxide donor to the normoxic culture promoted osteoclastogenesis. Furthermore, insulin-like growth factor 2 expression was significantly altered in both iNOS inhibition experiments and nitric oxide donor experiments. These data might provide clues to therapies for excessive osteoclastogenesis under several hypoxic pathologic conditions, including rheumatoid arthritis.
Topics: Animals; Bone Resorption; Cell Differentiation; Cell Hypoxia; Cells, Cultured; Enzyme Induction; Hypoxia; Male; Mice; Mice, Inbred C57BL; Nitric Oxide Synthase Type II; Osteoclasts; Osteogenesis; Oxygen; Signal Transduction; omega-N-Methylarginine
PubMed: 34560064
DOI: 10.1016/j.ajpath.2021.08.014 -
Experimental Animals Jan 2000The electrophysiological properties of cardiac muscle in KK/Ta mouse (hereafter referred to as KK mouse), an animal model of human non-insulin-dependent diabetes...
The electrophysiological properties of cardiac muscle in KK/Ta mouse (hereafter referred to as KK mouse), an animal model of human non-insulin-dependent diabetes mellitus, were investigated, and the findings compared with those obtained from a non-diabetic control mouse (C57BL/6J mouse; referred to as B6 mouse). The ages of the B6 mice were 23.9 +/- 5.4 weeks (n = 24) and those of the KK mice used were 25.7 +/- 10.8 weeks (n = 34). The KK mice had mild obesity, hyperglycemia and hyperinsulinemia. Ventricular muscles from both mice were examined by light microscopy. Partial myocardial fibrosis and filament disorder in the ventricular muscles were found only in the KK mice. The resting membrane potential of the ventricular muscle was less negative in the KK mice than in the control mice. The maximum rate of rise in the upstroke of the action potential was significantly decreased in the KK mice compared with that of the control mice. These suggest a decrease in a time-independent K+ current (IK1) in the KK mice. The duration of the action potential (APD) at all levels of repolarization was significantly longer in the KK mice than in the B6 mice. A blocker of transient outward current (I(to)), 4-aminopyridine, significantly prolonged the APD of the B6 mice, but failed to prolong it in the KK mice, suggesting that Ito in the diabetic mice is very small. A Ca2+ channel blocker, CoCl2, dramatically lengthened all levels of APD in both groups, suggesting that there is no difference between B6 mice and KK mice in L-type Ca2+ current via Ca2+ channels. These suggest the malfunction or deficiency of ionic channels which carry, at least Ito and IK1 in diabetic mice.
Topics: 4-Aminopyridine; Action Potentials; Aging; Animals; Blood Glucose; Calcium Channel Blockers; Calcium Channels, L-Type; Cobalt; Diabetes Mellitus, Type 2; Electric Conductivity; Electrophysiology; Heart Ventricles; Insulin; Male; Membrane Potentials; Mice; Mice, Inbred C57BL; Obesity; Pancreas; Potassium Channels
PubMed: 10803358
DOI: 10.1538/expanim.49.23 -
Psychopharmacology Jan 2024Clozapine N-oxide (CNO) has been developed as a ligand to selectively activate designer receptors exclusively activated by designer drugs (DREADDs). However, previous...
RATIONALE
Clozapine N-oxide (CNO) has been developed as a ligand to selectively activate designer receptors exclusively activated by designer drugs (DREADDs). However, previous studies have revealed that peripherally injected CNO is reverse-metabolized into clozapine, which, in addition to activating DREADDs, acts as an antagonist at various neurotransmitter receptors, suggesting potential off-target effects of CNO on animal physiology and behaviors. Recently, second-generation DREADD agonists compound 21 (C21) and JHU37160 (J60) have been developed, but their off-target effects are not fully understood.
OBJECTIVES
The present studies assessed the effect of novel DREADD ligands on reward-seeking behavior.
METHODS
We first tested the possible effect of acute i.p. injection of low-to-moderate (0.1, 0.3, 1, 3 mg/kg) of CNO, C21, and J60 on motivated reward-seeking behavior in wild-type mice. We then examined whether a high dose (10 mg/kg) of these drugs might be able to alter responding.
RESULTS
Low-to-moderate doses of all drugs and a high dose of CNO or C21 did not alter operant lick responding for a reward under a progressive ratio schedule of reinforcement, in which the number of operant lick responses to obtain a reward increases after each reward collection. However, high-dose J60 resulted in a total lack of responding that was later observed in an open field arena to be due to a sedative effect.
CONCLUSIONS
This study provides definitive evidence that commonly used doses of CNO, C21, and J60 have negligible off-target effects on motivated reward-seeking but urges caution when using high doses of J60 due to sedative effects.
Topics: Mice; Animals; Clozapine; Reward; Designer Drugs
PubMed: 37792024
DOI: 10.1007/s00213-023-06465-w -
Experimental Animals Jan 2001In the present study, we investigated the characteristics of the postrest contraction (PRC) in chronic diabetic ventricular muscle. We used WBN/Kob rats of 7-8 weeks as...
In the present study, we investigated the characteristics of the postrest contraction (PRC) in chronic diabetic ventricular muscle. We used WBN/Kob rats of 7-8 weeks as the spontaneously diabetic animal and Wistar rats of 7-8 weeks as the control. We found: (1) No significant differences were seen in the amplitude, the contracting speed, and the relaxing speed of electrically stimulated twitch tension between control and WBN/Kob rats. In addition, the relationship between amplitude of twitch tension and stimulus cycle lengths (0.2-5 sec) was very similar in both animals. (2) The ratios of the first twitch tension (T1) of PRC with various rest intervals (5-600 sec) to the steady-state tension (Tss) were significantly smaller in the diabetic rats than in the controls. (3) When the preparation was stimulated at shorter cycle lengths, the recovery process of PRC was separated into at least two components (fast and slow components). In the diabetic rats, the time constant (tau) of both components was significantly longer than in controls. (4) After caffeine (10(-3) M) treatment, tau of the fast component in the control rats became longer, whereas it remained unchanged in diabetic rats. These findings suggest a dysfunction of the intracellular calcium handling system in spontaneously diabetic heart that is likely to include impaired calcium sequestration and/or extrusion.
Topics: Animals; Blood Glucose; Body Weight; Caffeine; Calcium; Diabetes Mellitus; Electric Stimulation; Male; Myocardial Contraction; Papillary Muscles; Rats; Rats, Wistar; Sarcoplasmic Reticulum; Time Factors
PubMed: 11326420
DOI: 10.1538/expanim.50.19 -
The Japanese Journal of Physiology 1990Myoplasmic Ca2+ metabolism is reported to be impaired in diabetic rat heart. We studied the possibility that the ventricular muscles of diabetic guinea pig are prone to...
Myoplasmic Ca2+ metabolism is reported to be impaired in diabetic rat heart. We studied the possibility that the ventricular muscles of diabetic guinea pig are prone to develop delayed afterdepolarizations (DADs) and triggered activity (TA), because DADs and TA are believed to be a possible index of increased level of intracellular Ca2+ concentration. To establish an experimental diabetic model from the guinea pig, male animals were divided into four groups: 1) control group: intracardiac injection of citrate buffer; 2) IP group: intraperitoneal injection of streptozotocin (STZ, 200 mg/kg); 3) IC group: intracardiac injection of STZ; and 4) Ins-IC group: intracardiac injection of STZ after pretreatment with insulin (20 IU/kg). We found that: 1) only in the Ins-IC group was the fasting plasma glucose concentration (determined 40 days after STZ injection) significantly higher than in the control group; 2) oral glucose tolerance test performed 40 days after treatment also showed glucose intolerance in the Ins-IC group. These findings evinced the successful making of diabetic guinea pigs by an intracardiac one-shot injection of STZ during development of insulin-induced hypoglycemia. In vitro electrophysiological experiments were performed on ventricular papillary muscle from diabetic animals (Ins-IC group) by conventional glass microelectrode techniques. Transmembrane action potentials were elicited by pulse trains with various rates (2-5 Hz) and durations (10-30 stimuli) in the presence of ouabain (1 microM) and various Ca2+ concentrations (1.8-7.2 mM). The incidence of TA in the muscles from diabetic animals was significantly higher (chi 2-test, p less than 0.05) than that from controls. The findings gave evidence that Ca2+ homeostasis in the myocardium of diabetic guinea pigs is impaired, and this may be a cause of arrhythmia.
Topics: Action Potentials; Animals; Blood Glucose; Calcium; Diabetes Mellitus, Experimental; Electrophysiology; Guinea Pigs; Heart; Heart Ventricles; Male; Myocardium
PubMed: 2086986
DOI: 10.2170/jjphysiol.40.651