Competitive Interactions between Halothane and Isoflurane at the Carotid Body and TASK Channels

Background The degree to which different volatile anesthetics depress carotid body hypoxic response relates to their ability to activate TASK potassium channels. Most commonly, volatile anesthetic pairs act additively at their molecular targets. We examined whether this applied to carotid body TASK channels. Methods We studied halothane and isoflurane effects on hypoxia-evoked rise in intracellular calcium (Ca2+i, using the indicator Indo-1) in isolated neonatal rat glomus cells, and TASK single-channel activity (patch clamping) in native glomus cells and HEK293 cell line cells transiently expressing TASK-1. Results Halothane (5%) depressed glomus cell Ca2+i hypoxic response (mean ± SD, 94 ± 4% depression; P < 0.001 vs. control). Isoflurane (5%) had a less pronounced effect (53 ± 10% depression; P < 0.001 vs. halothane). A mix of 3% isoflurane/1.5% halothane depressed cell Ca2+i response (51 ± 17% depression) to a lesser degree than 1.5% halothane alone (79 ± 15%; P = 0.001), but similar to 3% isoflurane alone (44 ± 22%; P = 0.224), indicating subadditivity. Halothane and isoflurane increased glomus cell TASK-1/TASK-3 activity, but mixes had a lesser effect than that seen with halothane alone: 4% halothane/4% isoflurane yielded channel open probabilities 127 ± 55% above control, versus 226 ± 12% for 4% halothane alone (P = 0.009). Finally, in HEK293 cell line cells, progressively adding isoflurane (1.5 to 5%) to halothane (2.5%) reduced TASK-1 channel activity from 120 ± 38% above control, to 88 ± 48% (P = 0.034). Conclusions In all three experimental models, the effects of isoflurane and halothane combinations were quantitatively consistent with the modeling of weak and strong agonists competing at a common receptor on the TASK channel. Editor’s Perspective What We Already Know about This Topic What This Article Tells Us That Is New

Download Full-text

Lemon Juice Mediated Synthesis of 3-Substituted Quinazolin-4(3H)-Ones and their Pharmacological Evaluation

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520619666190723151909 ◽

2020 ◽

Vol 19 (16) ◽

pp. 2001-2009 ◽

Cited By ~ 1

Author(s):

Malavattu G. Prasad ◽

C. Vijaya Lakshmi ◽

Naresh K. Katari ◽

Sreekantha B. Jonnalagadda ◽

Manojit Pal

Keyword(s):

Cell Line ◽

Cell Lines ◽

Hek293 Cell ◽

Glyoxylic Acid ◽

Lemon Juice ◽

Cytotoxic Agents ◽

Diverse Range ◽

Hek293 Cell Line ◽

Three Component Reaction ◽

Mcf 7

Background: Compounds containing the quinazoline-4(3H)-one framework constitute an important class of fused N-heterocycles that are found in more than 200 naturally occurring alkaloids. These compounds also show a diverse range of pharmacological activities including antitumor properties. This prompted us to explore a series of quinazolin-4-(3H)-one derivatives having no substituent at C-2 as potential cytotoxic agents. Objective: The objective of this study was to synthesize and evaluate 3-substituted quinazolin-4(3H)-one derivatives for their potential cytotoxic properties. Methods: A convenient method has been developed for the rapid synthesis of this class of compounds under a mild and non-hazardous reaction condition in good yields. The methodology involved a three-component reaction employing isatoic anhydride, amines and glyoxylic acid as reactants in the presence of lemon juice in PEG- 400 at room temperature (25-30ºC) under ultrasound irradiation. All the synthesized compounds were screened via an MTT assay for their potential cytotoxic properties in vitro using the cancerous cell lines e.g. A549, A2780, HepG2, K562, MCF-7 and HCT-116 and a non-cancerous HEK293 cell line. Results: Several compounds such as 3a, 3b, 3d, 3e and 3f showed promising growth inhibition against these cancer cell lines but no significant effects on HEK293 cell line. The IC50 values of these compounds were comparable to doxorubicin whereas 3f significantly induced apoptosis in MCF-7 cells that also was comparable to doxorubicin. Conclusion: An ultrasound-assisted MCR facilitated by lemon juice has been developed to synthesize 3- substituted quinazolin-4(3H)-one derivatives that could act as potential anticancer agents.

Download Full-text

Affecting HEK293 Cell Growth and Production Performance by Modifying the Expression of Specific Genes

Cells ◽

10.3390/cells10071667 ◽

2021 ◽

Vol 10 (7) ◽

pp. 1667

Author(s):

Laura Abaandou ◽

David Quan ◽

Joseph Shiloach

Keyword(s):

Cell Line ◽

Cell Lines ◽

Hek293 Cell ◽

Chinese Hamster ◽

Production Performance ◽

Cellular Processes ◽

Hek293 Cell Line ◽

Global Engineering ◽

Genomic Tools ◽

Serum Free Suspension

The HEK293 cell line has earned its place as a producer of biotherapeutics. In addition to its ease of growth in serum-free suspension culture and its amenability to transfection, this cell line’s most important attribute is its human origin, which makes it suitable to produce biologics intended for human use. At the present time, the growth and production properties of the HEK293 cell line are inferior to those of non-human cell lines, such as the Chinese hamster ovary (CHO) and the murine myeloma NSO cell lines. However, the modification of genes involved in cellular processes, such as cell proliferation, apoptosis, metabolism, glycosylation, secretion, and protein folding, in addition to bioprocess, media, and vector optimization, have greatly improved the performance of this cell line. This review provides a comprehensive summary of important achievements in HEK293 cell line engineering and on the global engineering approaches and functional genomic tools that have been employed to identify relevant genes for targeted engineering.

Download Full-text

An anti-apoptotic HEK293 cell line provides a robust and high titer platform for transient protein expression in bioreactors

mAbs ◽

10.1080/19420862.2019.1598230 ◽

2019 ◽

Vol 11 (5) ◽

pp. 977-986 ◽

Cited By ~ 2

Author(s):

Tia A Arena ◽

Bernice Chou ◽

Peter D. Harms ◽

Athena W. Wong

Keyword(s):

Protein Expression ◽

Cell Line ◽

Hek293 Cell ◽

High Titer ◽

Hek293 Cell Line ◽

Transient Protein Expression

Download Full-text

Carotid body chemosensory responses in mice deficient of TASK channels

The Journal of General Physiology ◽

10.1085/jgp.200910302 ◽

2010 ◽

Vol 135 (4) ◽

pp. 379-392 ◽

Cited By ~ 61

Author(s):

Patricia Ortega-Sáenz ◽

Konstantin L. Levitsky ◽

María T. Marcos-Almaraz ◽

Victoria Bonilla-Henao ◽

Alberto Pascual ◽

...

Keyword(s):

Carotid Body ◽

Sensory Transduction ◽

Resting Potential ◽

Cell Phenotype ◽

Null Mouse ◽

Wild Type ◽

Glomus Cell ◽

Secretory Rate ◽

K Channels ◽

Glomus Cells

Background K+ channels of the TASK family are believed to participate in sensory transduction by chemoreceptor (glomus) cells of the carotid body (CB). However, studies on the systemic CB-mediated ventilatory response to hypoxia and hypercapnia in TASK1- and/or TASK3-deficient mice have yielded conflicting results. We have characterized the glomus cell phenotype of TASK-null mice and studied the responses of individual cells to hypoxia and other chemical stimuli. CB morphology and glomus cell size were normal in wild-type as well as in TASK1−/− or double TASK1/3−/− mice. Patch-clamped TASK1/3-null glomus cells had significantly higher membrane resistance and less hyperpolarized resting potential than their wild-type counterpart. These electrical parameters were practically normal in TASK1−/− cells. Sensitivity of background currents to changes of extracellular pH was drastically diminished in TASK1/3-null cells. In contrast with these observations, responsiveness to hypoxia or hypercapnia of either TASK1−/− or double TASK1/3−/− cells, as estimated by the amperometric measurement of catecholamine release, was apparently normal. TASK1/3 knockout cells showed an enhanced secretory rate in basal (normoxic) conditions compatible with their increased excitability. Responsiveness to hypoxia of TASK1/3-null cells was maintained after pharmacological blockade of maxi-K+ channels. These data in the TASK-null mouse model indicate that TASK3 channels contribute to the background K+ current in glomus cells and to their sensitivity to external pH. They also suggest that, although TASK1 channels might be dispensable for O2/CO2 sensing in mouse CB cells, TASK3 channels (or TASK1/3 heteromers) could mediate hypoxic depolarization of normal glomus cells. The ability of TASK1/3−/− glomus cells to maintain a powerful response to hypoxia even after blockade of maxi-K+ channels, suggests the existence of multiple sensor and/or effector mechanisms, which could confer upon the cells a high adaptability to maintain their chemosensory function.

Download Full-text

Comparative Genomics, Infectivity and Cytopathogenicity of American Isolates of Zika Virus that Developed Persistent Infections in Human Embryonic Kidney (HEK293) Cells

International Journal of Molecular Sciences ◽

10.3390/ijms20123035 ◽

2019 ◽

Vol 20 (12) ◽

pp. 3035 ◽

Cited By ~ 4

Author(s):

Hebing Liu ◽

Hsiao-Mei Liao ◽

Bingjie Li ◽

Shien Tsai ◽

Guo-Chiuan Hung ◽

...

Keyword(s):

Cell Line ◽

Zika Virus ◽

Hek293 Cell ◽

Clonal Selection ◽

Gene Mutations ◽

Host Cells ◽

Hek293 Cells ◽

Specific Gene ◽

Human Embryonic Kidney ◽

Hek293 Cell Line

Zika virus (ZIKV) transmission can cause serious fetal neurological abnormalities. ZIKV persistence in various human cells and tissues can serve as infectious reservoirs and post serious threats to public health. The human embryonic kidney (HEK293) cell line with known neuronal developmental properties was readily infected by ZIKV in a strain-dependent fashion. Significant cytopathic effect in HEK293 cells infected by the prototype MR 766 strain of ZIKV resulted in complete loss of cells, while small numbers of HEK293 cells infected by contemporary ZIKV isolates (PRV or FLR strain) continued to survive and regrow to confluency in the culture around two months after initial infection. Most, if not all, of the cells in the two resulting persistently ZIKV-infected HEK293 cell lines tested positive for ZIKV antigen. Compared to HEK293 control cells, the persistently ZIKV-infected HEK293 cells had slower growth rates with some cells undergoing apoptosis in culture. The “persistent ZIKVs” produced constitutively by both PRV and FLR strains ZIKV-infected HEK293 cells had significantly attenuated cell infectivity and/or cytopathogenicity. Comparative genome sequence analyses between the persistent ZIKVs and the original inoculum ZIKVs showed no clonal selection with specific gene mutations in the prolonged process of establishing persistently PRV strain ZIKV-infected HEK293 cells; while selection of ZIKV subclones with mutations in the envelope, protein pr and multiple NS genes was evident in developing persistently FLR strain ZIKV-infected HEK293 cell line. Our study provides molecular insights into the complex interplays of ZIKV and human host cells in establishing ZIKV persistence.

Download Full-text

Effects of PAX2 expression in a human fetal kidney (HEK293) cell line

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research ◽

10.1016/s0167-4889(97)00119-5 ◽

1998 ◽

Vol 1401 (1) ◽

pp. 53-62 ◽

Cited By ~ 27

Author(s):

Elena Torban ◽

Paul R Goodyer

Keyword(s):

Cell Line ◽

Hek293 Cell ◽

Fetal Kidney ◽

Hek293 Cell Line ◽

Pax2 Expression ◽

Human Fetal Kidney

Download Full-text

Transduction mechanisms in carotid body: glomus cells, putative neurotransmitters, and nerve endings

AJP Cell Physiology ◽

10.1152/ajpcell.1980.239.5.c135 ◽

1980 ◽

Vol 239 (5) ◽

pp. C135-C152 ◽

Cited By ~ 77

Author(s):

C. Eyzaguirre ◽

S. J. Fidone

Keyword(s):

Carotid Body ◽

Conclusive Evidence ◽

Sensory Transduction ◽

Nerve Endings ◽

High Pco2 ◽

Type I ◽

Glomus Cell ◽

Impulse Frequency ◽

Glomus Cells ◽

Carotid Body Chemoreceptors

Carotid body chemoreceptors are activated by low PO2, high PCO2, acidity, increased temperature, and tonicity. These receptors are important in homeostasis and mediate their reflex effects on the CNS through sensory discharges of the carotid (sinus) nerve. The receptor complex is formed by glomus (type I) cells and carotid nerve endings, which, morphologically, appear to form a sensory synapse. The junction between glomus cells and nerve endings is enveloped by processes of sustentacular (type II) cells. The mechanisms of chemoreceptor transduction are complex; there is no agreement about the identity of the primary receptor element (glomus cell or nerve terminal) or what mechanisms are responsible for the onset of the sensory discharge in the carotid nerve. There is increasing evidence that integrity of the glomus cell is essential for normal transduction and that the receptor synapse described by morphologists may be functionally active. There is no conclusive evidence, however, that the glomus cell is the primary site of sensory transduction. Stimuli act on the glomus cell to release "transmitter" and/or "modulator" substances; but it is unknown if the released chemicals are directly responsible for the accompanying change in sensory impulse frequency or merely modify an already ongoing discharge. Interactions between glomus cells and nerves may be complicated enough to make it very difficult to resolve this question.

Download Full-text

Abstract 956: Acid-Sensing Ion Channels (ASICs) Contribute to Transduction of Extracellular Acidosis in Rat Carotid Body Glomus Cells

Circulation ◽

10.1161/circ.116.suppl_16.ii_189 ◽

2007 ◽

Vol 116 (suppl_16) ◽

Author(s):

Zhi-Yong Tan ◽

Yongjun Lu ◽

Carol A Whiteis ◽

Christopher J Benson ◽

Mark W Chapleau ◽

...

Keyword(s):

Ion Channels ◽

Carotid Body ◽

Bathing Solution ◽

Ph Sensing ◽

Pipette Solution ◽

Glomus Cells ◽

Extracellular Acidosis ◽

Task Channels ◽

Carotid Body Chemoreceptors ◽

Carotid Body Glomus Cells

The molecular mechanism of pH sensing by chemoreceptors is not clear, although it had been proposed to be mediated by a drop in intracellular pH of carotid body glomus cells, which inhibits a K + current. Recently, pH-sensitive ion channels have been described in glomus cells that respond directly to extracellular acidosis. In this study, we investigated the possible molecular mechanisms of carotid body pH-sensing by recording the responses of glomus cells isolated from rat carotid body to rapid changes in extracellular pH using whole-cell patch-clamping technique. Extracellular acidosis evoked transient inward currents in glomus cells that were evident at pH 7.0 and half-activated (pH 50) at 6.3. The current had the characteristics of ASICs. It averaged 40.7±15.7 pA (n=5) at pH 5.0 and was blocked by the ASIC channel blocker amiloride (200 μm) to 2.5±1.6 pA. Na + free bathing solution eliminated the current and a Ca 2+ free buffer enhanced (P<0.05) the current at pH 6.0 from 18.5±2.2 to 86.0±12.5 pA (n=5). Enhancement of the current was also seen with the addition of lactate. In the current clamp mode extracellular acidosis evoked both a transient and sustained depolarization. The initial transient component at pH 6.0 averaged 18.2±2.6 mV and was blocked by amiloride to 2.1±2.1 mV supporting the contribution of ASICs. However, the sustained depolarization was not blocked by amiloride but was eliminated by removal of K + from the pipette solution which reduced significantly intracellular K + . This sustained depolarization was partially blocked by the TASK channels blockers anandamide (from 14.9±1.6 mV to 9.3±2.2 mV at pH 6.0, n=5) and quinidine (from 27.5±2.2 mV to 11.3±2.3 mV at pH 6.0, n=3). The results provide the first evidence that ASICs may contribute to chemotransduction of low pH by carotid body chemoreceptors, and that extracellular acidosis directly activates carotid body chemoreceptors through both ASIC and TASK channels.

Download Full-text