scholarly journals Formation of gap junctions by treatment in vitro with potassium conductance blockers.

1978 ◽  
Vol 78 (2) ◽  
pp. 338-348 ◽  
Author(s):  
M S Kannan ◽  
E E Daniel
Keyword(s):  
Smooth Muscle ◽  
Gap Junctions ◽  
De Novo ◽  
Potassium Conductance ◽  
Cell Coupling ◽  
Thin Sections ◽  
Rapid Induction ◽  
Space Constant ◽  
Spontaneous Contractions

Gap junctions were regularly seen in thin sections of canine tracheal smooth muscle incubated in vitro. Their number was increased in tissued exposed in vitro to either of two potassium conductance blockers, tetraethylammonium (TEA) and 4-aminopyridine (4-AP), and at the same time the muscles became mechanically active, with spontaneous contractions. The presence of gap junctions in this smooth muscle may provide one basis for cell-to-cell coupling, and their increase after TEA- and 4-AP-treatment could account for a decreased junctional resistance between cells, contributing to a longer space constant. However, an increase in gap junctions was not sufficient to change the behavior of trachealis smooth muscle from multiunit to single-unit type. Gap junctions in increased numbers persisted after washout of 4-AP, which caused inhibition of spontaneous contractions, and despite inhibition of the contractile effects of 4-AP by atropine. The rapid induction of gap junction formation was not dependent on de novo synthesis of protein. The fact that the number of gap junctions can be increased by chemical agents has important implications for control of their formation and provides a tool for analysis fo their role in cell-to-cell coupling.

scholarly journals Vitamin-A-induced mucous metaplasia. An in vitro system for modulating tight and gap junction differentiation.

1976 ◽  
Vol 68 (2) ◽  
pp. 173-188 ◽  
Author(s):  
P M Elias ◽  
D S Friend
Keyword(s):  
Vitamin A ◽  
Gap Junctions ◽  
Gap Junction ◽  
De Novo ◽  
Freeze Fracture ◽  
Controlled Study ◽  
Thin Sections ◽  
Tight Junction Formation ◽  
Mucous Metaplasia

Stratified squamous epithelia from 14-day chick embryo shank skin contain rare tight-junctional strands and only small gap junctions. Exposure of this tissue to retinoic acid (vitamin-A) (20 U/ml) in organ culture, however, induces mucous metaplasia, accompanied by tight-junction formation and gap-junction growth; untreated specimens continue to keratinize. To investigate sequential stages of junctional assembly and growth, we examined thin sections and freeze-fracture replicas at daily intervals for 3 days. During the metaplastic process, tight junctions assemble in midepidermal and upper regions, beginning on day 1 and becoming maximal on day 3. Two tight-junctional patterns could be tentatively identified as contributing to the emergence of fully formed zonulae occludentes: (a) the formation of individual ridges along the margins of gap junctions; (b) de novo generation of continuous ramifying strands by fusion of short strand segments and linear particulate aggregates near cellular apices. Gap junction enlargement, already maximal at day 1, occurs primarily three to four cell layers deep. Growth appears to occur by annexation of islands of 20-40 8.5-nm particles into larger lattices of islands separated by particle-free aisles. Eventually, a single gap junction may occupy much of the exposed membrane face in freeze-fractured tissue, but during apical migration of the cells such junctions disappear. The vitamin- A chick-skin system is presented as a responsive model for the controlled study of junction assembly.

Are gap junctions necessary for cell-to-cell coupling of smooth muscle?: an update

1992 ◽  
Vol 70 (4) ◽  
pp. 481-490 ◽  
Author(s):  
R. E. Garfield ◽  
G. Thilander ◽  
M. G. Blennerhassett ◽  
N. Sakai
Keyword(s):  
Smooth Muscle ◽  
Gap Junctions ◽  
Current Knowledge ◽  
Smooth Muscles ◽  
Cell Communication ◽  
Circular Muscle ◽  
Cell Coupling ◽  
And Function ◽  
Longitudinal Layer ◽  
Cell Cell

Earlier, it was questioned whether gap junctions (GJs) were necessary for cell–cell communication in smooth muscle, and GJs were not seen in some smooth muscles. We reexamined this question in the myometrium and in intestinal smooth muscle, in light of current knowledge of the presence and function of GJs. In the uterus, numerous studies show that an increase in GJ number is associated with the onset of delivery and is required for effective parturition. In all cases, this increase in GJ number and the changes in uterine contractility were correlated with increased electrical and metabolic coupling. Evidence for the much smaller, but detectable, degree of electrical coupling in the preterm uterus is explained by the small (but again detectable) number of GJs present. In the intestine, GJs are readily detected in the circular muscle layer but have not been described in the adjacent longitudinal layer. While our immunohistochemical studies failed to detect GJs in the longitudinal layer, this may not be adequate to prove their absence. Therefore, current knowledge of GJ number and function is adequate to explain cell–cell coupling in the uterus. Although it remains uncertain whether GJs are absent from the longitudinal muscle of the intestine, there is no definitive evidence that cell–cell coupling can occur by means other than GJs.Key words: gap junctions, myometrium, connexins, smooth muscle, cell communication.

Stretch-mediated visceral smooth muscle growth in vitro

1992 ◽  
Vol 262 (5) ◽  
pp. R895-R900
Author(s):  
O. M. Karim ◽  
K. Pienta ◽  
N. Seki ◽  
J. L. Mostwin
Keyword(s):  
Smooth Muscle ◽  
Dna Synthesis ◽  
De Novo ◽  
Specific Activity ◽  
Muscle Growth ◽  
Mechanical Stimulus ◽  
Trophic Factors ◽  
Taenia Coli ◽  
Visceral Smooth Muscle

An in vitro model of smooth muscle stretch was developed to study mechanical stimulus as a possible mediator of visceral smooth muscle growth and differences in the growth response of smooth muscle from young and old animals. De novo DNA synthesis as measured by the aphidicolin-sensitive specific activity of DNA was used as an index of cell growth. Compared with old tissue, the rate of aphidicolin-sensitive DNA synthesis in smooth muscle from young animals was 3-5 and 1.5-2 times greater in bladder and taenia coli, respectively. Stretch of bladder muscle and taenia coli strips from young animals for 6 h increased the aphidicolin-sensitive specific activity of DNA 3-fold (P less than 0.01) and 1.5-fold (P less than 0.01), respectively. Tissue from old animals, however, under the same conditions increased the rate of aphidicolin-resistant DNA synthesis, possibly implying DNA repair. Autoradiography showed only labeled myocyte nuclei. These results indicate that homeostatic mechanisms modulating myocyte growth in visceral smooth muscle can respond to mechanical stimulus in the absence of other trophic factors.

Neural regulation of in vitro giant contractions in the rat colon

2001 ◽  
Vol 281 (1) ◽  
pp. G275-G282 ◽  
Author(s):  
Asensio Gonzalez ◽  
Sushil K. Sarna
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Sch 23390 ◽  
Circular Muscle ◽  
Electrical Field Stimulation ◽  
Muscle Cells ◽  
Enteric Neurons ◽  
Rat Colon ◽  
Spontaneous Contractions

The rat middle colon spontaneously generates regularly occurring giant contractions (GCs) in vitro. We investigated the neurohumoral and intracellular regulation of these contractions in a standard muscle bath. cGMP content was measured in strips and single smooth muscle cells. The circular muscle strips generated spontaneous GCs. Their amplitude and frequency were significantly increased by tetrodotoxin (TTX), ω-conotoxin, N ω-nitro-l-arginine (l-NNA), and the dopamine D1 receptor antagonist Sch-23390. The GCs were unaffected by hexamethonium, atropine, and antagonists of serotonergic (5-HT1–4), histaminergic (H1–2), and tachykininergic (NK1–2) receptors but enhanced by NK3receptor antagonism. The guanylate cyclase inhibitor 1H-[1,2,4]oxidiazolo[4,3-a]quinoxalin-1-one (ODQ) also enhanced GCs to the same extent as TTX and l-NNA, and each of the three agents prevented the effects of the others. GCs were abolished by electrical field stimulation, S-nitroso- N-acetyl-penicillamine, and 8-bromo-cGMP. BAY-K-8644 and apamin enhanced the GCs, but they were abolished by D-600. Basal cGMP content in strips was decreased by TTX,l-NNA, or ODQ, but these treatments had no effect on cGMP content of enzymatically dissociated single smooth muscle cells. We conclude that spontaneous contractions in the rat colonic muscle strips are not generated by cholinergic, serotonergic, or histaminergic input. Constitutive release of nitric oxide from enteric neurons sustains cGMP synthesis in the colonic smooth muscle to suppress spontaneous in vitro GCs.

Possible role of gap junctions in activation of myometrium during parturition

1978 ◽  
Vol 235 (5) ◽  
pp. C168-C179 ◽  
Author(s):  
R. E. Garfield ◽  
S. M. Sims ◽  
M. S. Kannan ◽  
E. E. Daniel
Keyword(s):  
Smooth Muscle ◽  
Gap Junctions ◽  
Freeze Fracture ◽  
Normal Pregnancy ◽  
Pregnant Rats ◽  
Junction Formation ◽  
Synchronous Contraction ◽  
Do So

Gap junctions between smooth muscle cells of the myometrium of pregnant rats were found only immediately prior to, during and immediately after parturition by quantitative thin-section and freeze-fracture microscopy. Ovariectomy of 16- to 17-days-pregnant rats resulted in premature termination of pregnancy and the appearance of gap junctions. Methods that prolonged normal pregnancy in rats or maintained pregnancy in ovariectomized animals (progesterone treatment) prevented the appearance of gap junctions. Gap junctions formed in tissues incubated for 24--96 h in vitro without any hormonal influence. We propose that gap junctions are essential for normal labor and delivery for synchronous contraction of the muscle of the uterus. We present a model for control of parturition that may apply to other animals including humans. The model proposes: 1) the possible roles progesterone, prostaglandins, or estrogens may play in initiating gap-junction formation; 2) that the formation of gap junctions is a necessary step in activation of the myometrium leading to labor; and 3) that agents used to stimulate or inhibit labor may do so by affecting gap junctions.

Extravascular sources of lung angiotensin peptide synthesis in idiopathic pulmonary fibrosis

2006 ◽  
Vol 291 (5) ◽  
pp. L887-L895 ◽  
Author(s):  
Xiaopeng Li ◽  
Maria Molina-Molina ◽  
Amal Abdul-Hafez ◽  
Jose Ramirez ◽  
Anna Serrano-Mollar ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Idiopathic Pulmonary Fibrosis ◽  
Pulmonary Fibrosis ◽  
Human Lung ◽  
De Novo ◽  
Alveolar Epithelial Cells ◽  
Human Lung Cells ◽  
Normal Human

Previous work from this laboratory demonstrated de novo synthesis of angiotensin (ANG) peptides by apoptotic pulmonary alveolar epithelial cells (AEC) and by lung myofibroblasts in vitro and in bleomycin-treated rats. To determine whether these same cell types also synthesize ANG peptides de novo within the fibrotic human lung in situ, we subjected paraffin sections of normal and fibrotic (idiopathic pulmonary fibrosis, IPF) human lung to immunohistochemistry (IHC) and in situ hybridization to detect ANG peptides and angiotensinogen (AGT) mRNA. These were analyzed both alone and in combination with cell-specific markers of AEC [monoclonal antibody (MAb) MNF-116] and myofibroblasts [α-smooth muscle actin (α-SMA) MAb] and an in situ DNA end labeling (ISEL) method to detect apoptosis. In normal human lung, IHC detected AGT protein in smooth muscle underlying normal bronchi and vessels, but not elsewhere. Real-time RT-PCR and Western blotting revealed that AGT mRNA and protein were 21-fold and 3.6-fold more abundant, respectively, in IPF lung biopsies relative to biopsies of normal human lung (both P < 0.05). In IPF lung, both AGT protein and mRNA were detected in AEC that double-labeled with MAb MNF-116 and with ISEL, suggesting AGT expression by apoptotic epithelia in situ. AGT protein and mRNA also colocalized to myofibroblast foci detected by α-SMA MAb, but AGT mRNA was not detected in smooth muscle. These data are consistent with earlier data from isolated human lung cells in vitro and bleomycin-induced rat lung fibrosis models, and they suggest that apoptotic AEC and myofibroblasts constitute key sources of locally derived ANG peptides in the IPF lung.

In-vitro characterization of the pharmacological effects induced by (–)-α-bisabolol in rat smooth muscle preparations

2012 ◽  
Vol 90 (1) ◽  
pp. 23-35 ◽  
Author(s):  
Rodrigo J.B. de Siqueira ◽  
Walter B.S. Freire ◽  
Alfredo A. Vasconcelos-Silva ◽  
Patrícia A. Fonseca-Magalhães ◽  
Francisco J.B. Lima ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Urinary Bladder ◽  
Biologically Active ◽  
Pharmacological Effects ◽  
Bladder Tissue ◽  
In Vitro Characterization ◽  
Voltage Dependent ◽  
Spontaneous Contractions

The present study deals with the pharmacological effects of the sesquiterpene alcohol (–)-α-bisabolol on various smooth-muscle preparations from rats. Under resting tonus, (–)-α-bisabolol (30–300 µmol/L) relaxed duodenal strips, whereas it showed biphasic effects in other preparations, contracting endothelium-intact aortic rings and urinary bladder strips, and relaxing these tissues at higher concentrations (600–1000 µmol/L). In preparations precontracted either electromechanically (by 60 mmol/L K+) or pharmacomechanically (by phenylephrine or carbachol), (–)-α-bisabolol showed only relaxing properties. The pharmacological potency of (–)-α-bisabolol was variable, being higher in mesenteric vessels, whereas it exerted relaxing activity with a lesser potency on tracheal or colonic tissues. In tissues possessing spontaneous activity, (–)-α-bisabolol completely decreased spontaneous contractions in duodenum, whereas it increased their amplitude in urinary bladder tissue. Administered in vivo, (–)-α-bisabolol attenuated the increased responses of carbachol in tracheal rings of ovalbumin-sensitized rats challenged with ovalbumin, but was without effect in the decreased responsiveness of urinary bladder strips in mice treated with ifosfamide. In summary, (–)-α-bisabolol is biologically active in smooth muscle. In some tissues, (–)-α-bisabolol preferentially relaxed contractions induced electromechanically, especially in tracheal smooth muscle. The findings from tracheal rings reveal that (–)-α-bisabolol may be an inhibitor of voltage-dependent Ca2+ channels.

scholarly journals Spontaneous and evoked contractions are regulated by PKC-mediated signaling in detrusor smooth muscle: involvement of BK channels

2013 ◽  
Vol 304 (5) ◽  
pp. F451-F462 ◽  
Author(s):  
Joseph A. Hypolite ◽  
Qi Lei ◽  
Shaohua Chang ◽  
Stephen A. Zderic ◽  
Stephan Butler ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Outlet Obstruction ◽  
Bk Channels ◽  
Bk Channel ◽  
Quiescent State ◽  
Detrusor Smooth Muscle ◽  
Muscle Involvement ◽  
Normal Bladder ◽  
Spontaneous Contractions

Protein kinase C (PKC) and large conductance Ca2+-activated potassium channels (BK) are downregulated in the detrusor smooth muscle (DSM) in partial bladder outlet obstruction (PBOO). DSM from these bladders display increased spontaneous activity. This study examines the involvement of PKC in the regulation of spontaneous and evoked DSM contractions and whether pharmacologic inhibition of PKC in normal DSM contributes to increased detrusor excitability. Results indicate the PKC inhibitor bisindolylmaleimide 1 (Bim-1) prevented a decline in the amplitude of spontaneous DSM contractions over time in vitro, and these contractions persist in the presence of tetrodotoxin. Bim-1 also reduced the basal DSM tone, and the ability to maintain force in response to electrical field stimulation, but did not affect maximum contraction. The PKC activator phorbol-12,13-dibutyrate (PDBu) significantly reduced the amplitude and increased the frequency of spontaneous contractions at low concentrations (10 nM), while causing an increase in force at higher concentrations (1 μM). Preincubation of DSM strips with iberiotoxin prevented the inhibition of spontaneous contractions by PDBu. The BK channel openers isopimaric acid and NS1619 reduced the Bim-1-induced enhancement of spontaneous contractions in DSM strips. Our data suggest that PKC has a biphasic activation profile in the DSM and that it may play an important role in maintaining the quiescent state of the normal bladder during storage through the effects on BK channel, while helping to maintain force required for bladder emptying. The data also suggest that PKC dysfunction, as seen in PBOO, contributes to detrusor overactivity.

scholarly journals Inverse Expression of Peroxisomes and Connexin-43 in the Granulosa Cells of the Quail Follicle

2000 ◽  
Vol 48 (2) ◽  
pp. 167-177 ◽  
Author(s):  
Stefano Farioli-Vecchioli ◽  
Stefaan Raes ◽  
Marc Espeel ◽  
Frank Roels ◽  
Katharina D'Herde
Keyword(s):  
Gap Junctions ◽  
Connexin 43 ◽  
Maturation Stage ◽  
Cell Coupling ◽  
Peroxisomal Disorders ◽  
Follicular Maturation ◽  
Marker Proteins ◽  
Disc Region ◽  
Cx 43

Studying the regulation of peroxisome (Px) expression could improve our understanding of human peroxisomal disorders. The granulosa of the largest preovulatory quail follicles proved to be a relevant model because (a) Px expression changes according to the follicular maturation stage and (b) Px expression varies regionally according to the distance of the granulosa relative to the germinal disc region containing the female gamete (oocyte). The question was asked whether Px expression is related to the extent of metabolic cell coupling and whether zonal Px variation is causally related to oocytal factors. This was evaluated by the presence of catalase and Cx-43 (marker proteins for peroxisomes and gap junctions, respectively) and by in vitro experiments with granulosa explants. The data obtained show that the expression of Cx-43 and Px is inversely correlated both temporally and spatially. Uncoupling of gap junctions results in an upregulation of α-catalase immunofluorescence. This is in agreement with reports that gap junctions are often negatively affected by Px proliferators. The zonal gradient in Px expression appears to be imposed by the oocyte, as is the case for steroidogenesis and proliferative capacity in the granulosa epithelium.

scholarly journals Discovery of an evolutionarily conserved smooth muscle cell-specific lncRNA CARMN

2020 ◽  
Author(s):  
Kunzhe Dong ◽  
Jian Shen ◽  
Xiangqin He ◽  
Liang Wang ◽  
Guoqing Hu ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
De Novo ◽  
Gene Expression Signature ◽  
Vascular Diseases ◽  
Cardiac Differentiation ◽  
Rna Seq ◽  
Specific Expression ◽  
Contractile Phenotype ◽  
Evolutionarily Conserved

AbstractDifferentiated vascular smooth muscle cells (VSMCs) are critical in maintaining vascular homeostasis by expressing a unique repertoire of contractile genes. Despite the well-defined coding transcriptome in differentiated VSMCs, little is known about the non-coding gene expression signature. Herein, by de novo analyzing publicly available RNA-seq and single cell RNA-seq datasets generated from different tissues and cell types, we unambiguously identified CARMN (CARdiac Mesoderm Enhancer-associated Non-coding RNA) as an evolutionarily conserved, SMC-specific lncRNA. CARMN was initially annotated as the host gene of MIR143/145 cluster and recently reported to play roles in cardiac differentiation. Here, we generated a Carmn GFP knock-in reporter mouse model and confirmed its specific expression in SMCs in vivo. In addition, we found Carmn is transcribed independently from Mir143/145 and only expressed transiently in embryonic cardiomyocytes and thereafter becomes restricted to adult SMCs in both human and mouse. Furthermore, we demonstrated that CARMN expression is not only dramatically decreased in human vascular diseases but functionally critical in maintaining VSMC contractile phenotype in vitro. In conclusion, we provided the first evidence showing that CARMN is an evolutionarily conserved SMC-specific lncRNA, down-regulated in different human vascular diseases, and a key lncRNA for maintaining SMC contractile phenotype.

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