Zebrafish or Danio rerio is an established model organism for studying the genetic, neuronal and behavioral bases of diseases and for toxicology and drug screening. The embryonic and larval stages of ...zebrafish have been used extensively in fundamental and applied research due to advantages offered such as body transparency, small size, low cost of cultivation and high genetic homology with humans. However, the manual experimental methods used for handling and investigating this organism are limited due to their low throughput, labor intensiveness and inaccuracy in delivering external stimuli to the zebrafish while quantifying various neuronal and behavioral responses. Microfluidic and lab-on-a-chip devices have emerged as ideal technologies to overcome these challenges. In this review paper, the current microfluidic approaches for investigation of behavior and neurobiology of zebrafish at embryonic and larval stages will be reviewed. Our focus will be to provide an overview of the microfluidic methods used to manipulate (deliver and orient), immobilize and expose or inject zebrafish embryos or larvae, followed by quantification of their responses in terms of neuron activities and movement. We will also provide our opinion in terms of the direction that the field of zebrafish microfluidics is heading toward in the area of biomedical engineering.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Microalgae are viable sources of biological compounds for biodiesel production. In this study, effects of various types of nitrogen sources and nutrients concentrations in the growth medium and ...different LED light wavelengths and intensities on biomass production of green algae Chlorella vulgaris were investigated. Warm white light with 80 µmol/m²/sec light intensity was determined as the optimal light for biomass production. The results indicated that microalgae growth with urea as nitrogen source was higher than that of other nitrogen sources such as sodium nitrate, ammonium carbonate and ammonium chloride. Maximum biomass concentration (1.37 g/L) was obtained under the following media compositions: urea 0.25 g/L, K₂HPO₄0.04 g/L, MgSO₄· 7H₂O 0.06 g/L, and ammonium ferric citrate 0.01 g/L. Microalgae growth data under the different light wavelengths and intensities were fitted with a mathematical model.
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•A power-law model including the effect of Brownian motion and thermophoresis is used.•A numerical scheme is employed to solve the highly non-linear equations.•The convective boundary ...condition is considered and studied for the stretching wall.•We model three kinds of nanofluids: pseudoplastic, Newtonian and dilatant nanofluids.•The effects of Brownian motion and thermophoresis are examined.
In this article, unsteady boundary layer flow of a power-law nanofluid over a stretching surface with a convective boundary condition is investigated numerically. A power-law model that incorporates the effects of Brownian motion and thermophoresis is used for non-Newtonian nanofluids. A set of similarity transformation is used to reduce Navier–Stokes, energy and nanoparticles concentration equations to a set of nonlinear ordinary differential equations which are then solved numerically by using a fourth order Runge–Kutta scheme coupled with a conventional shooting procedure. The effects of unsteadiness, suction/injection parameters, the generalized Prandtl and Lewis numbers and convective parameter on skin friction coefficient and reduced Nusselt number are investigated. Comparison with previously published work is performed and excellent agreement is observed for the limited case of existing literature. Numerical results show that dimensionless nanofluid concentration increases with the unsteadiness parameter whereas the values of dimensionless velocity and temperature decrease with it. Also, it is found that the effects of the unsteadiness parameter on the velocity boundary layer thickness are more pronounced compared to the concentration and temperature boundary layer thicknesses.
•A device to study the electric response of semi-mobile zebrafish larva in detail.•Zebrafish larvae sense the electric field (EF) direction and respond differently.•Voltage drop across larva’s body ...affects their response duration and tail frequency.•Exposure to multiple EF pulses result in decay in zebrafish larva’s response.•Two studies show that zebrafish larvae show habituation to EF pulses in the device.
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We previously showed that electric current induces zebrafish larvae to move towards the anode pole along a microchannel. For a larva with a fixed head and a moving tail, we observed that the response to electricity depended on the current magnitude. The effects of electric signal direction, voltage magnitude and habituation to repeated exposures to electric pulses were not characterized. Here, this knowledge gap was addressed by exploiting them in a microfluidic device with a head-trap to immobilize a zebrafish larva and a downstream chamber for tail movement phenotypic characterization based on response duration (RD) and tail beat frequency (TBF). We first assessed larvae’s response to electric current direction (at 3 µA) and voltage magnitude. Changing the current direction significantly altered the RD and TBF with long and low-frequency responses seen when the anode was positioned at the larvae’s tail. The electric voltage had a significant effect on larvae’s locomotion with long RD and low TBF observed at 5.6 V in the range of 1.3–9 V. We also demonstrated that the zebrafish locomotor response to repeated 3 µA current pulses diminished with dependency on the interstimulus interval. However, the diminished response was fully recovered after a 5-min resting period or introduction of a novel light stimulus (i.e., habituation-dishabituation strategy). Therefore, electric response suppression in zebrafish was attributed to the habituation as a form of non-associative learning. Our microfluidic platform has a broad application potential in behavioral neuroscience to study cognitive phenotypes, fundamental studies on the biological roots of electric response, and in pharmacological screening.
In this paper, we used DC electrophoretic force and electrical resistance measurement between two microwires in a microchannel to respectively extract and detect 1-10 μm polystyrene microplastics in ...water at 5-100 ppm concentration range.
Ex situ
microplastic detection methods like visual recognition and optical spectroscopy are expensive, time-consuming, and labor-intensive, highlighting the need for on-site rapid detection and quantification technologies. While micro-electro-fluidic devices have been introduced, they require advanced microfabrication and expensive signal processing instruments, while being prone to blockage with larger microplastics, limiting their point-of-need applications. We developed a simple wide PDMS channel equipped with two copper microwire electrodes, orthogonal to the flow direction and connected to a DC sourcemeter, for electrical resistance-based sensing of polystyrene microplastics in water. Electrical resistance between the two microwires was associated with microplastics' size and concentration, following their electrophoretic accumulation around the positive electrode. A positive correlation was found between microplastic concentration and reduction in the normalized resistance. Reducing the flow rate strengthened the extraction and detection of larger microplastics. In the future, the sensor will be tested for other types of microplastics in real samples and can be integrated into a handheld device for low cost and on-site microplastic detection.
DC electric field between two microwires in a straight microchannel was employed to electrophoretically accumulate and detect microplastics in water.
Abstract
Pannexin 1 (Panx1) forms ATP‐permeable membrane channels that play roles in purinergic signaling in the nervous system. A link between Panx1 activity and neurodegenerative disorders ...including Parkinson's disease (PD) has been suggested, but experimental evidence is limited. Here, a zebrafish model of PD was produced by exposing
panx1a
+/+
and
panx1a
−/−
zebrafish larvae to 6‐hydroxydopamine (6‐OHDA). Electrical stimulation in a microfluidic chip and quantitative real‐time‐qPCR of zebrafish larvae tested the role of Panx1 in both pathological and normal conditions. After 72‐h treatment with 6‐OHDA, the electric‐induced locomotor activity of 5 days post fertilization (5dpf)
panx1a
+/+
larvae were reduced, while the stimulus did not affect locomotor activity of age‐matched
panx1a
−/−
larvae. A RT‐qPCR analysis showed an increase in the expression of genes that are functionally related to dopaminergic signaling, like the tyrosine hydroxylase
(th2)
and the leucine‐rich repeat kinase 2
(lrrk2)
. Extending the 6‐OHDA treatment duration to 120 h caused a significant reduction in the locomotor response of 7dpf
panx1a
−/−
larvae compared to the untreated
panx1a
−/−
group. The RT‐qPCR data showed a reduced expression of dopaminergic signaling genes in both genotypes. It was concluded that the absence of Panx1a channels compromised dopaminergic signaling in 6‐OHDA‐treated zebrafish larvae and that the increase in the expression of dopaminergic genes was transient, most likely due to a compensatory upregulation. We propose that zebrafish Panx1a models offer opportunities to shed light on PD's physiological and molecular basis. Panx1a might play a role on the progression of PD, and therefore deserves further investigation.
Behavioral responses of zebrafish larvae to environmental cues are important functional readouts that should be evoked on-demand and studied phenotypically in behavioral, genetical and developmental ...investigations. Very recently, it was shown that zebrafish larvae execute a voluntary and oriented movement toward the positive electrode of an electric field along a microchannel. Phenotypic characterization of this response was not feasible due to larva's rapid movement along the channel. To overcome this challenge, a microfluidic device was introduced to partially immobilize the larva's head while leaving its mid-body and tail unrestrained in a chamber to image motor behaviors in response to electric stimulation, hence achieving quantitative phenotyping of the electrically evoked movement in zebrafish larvae. The effect of electric current on the tail-beat frequency and response duration of 5-7 days postfertilization zebrafish larvae was studied. Investigations were also performed on zebrafish exposed to neurotoxin 6-hydroxydopamine and larvae carrying a pannexin1a (panx1a) gene knockout, as a proof of principle applications to demonstrate on-demand movement behavior screening in chemical and mutant assays. We demonstrated for the first time that 6-hydroxydopamine leads to electric response impairment, levodopa treatment rescues the response and panx1a is involved in the electrically evoked movement of zebrafish larvae. We envision that our technique is broadly applicable as a screening tool to quantitatively examine zebrafish larvae's movements in response to physical and chemical stimulations in investigations of Parkinson's and other neurodegenerative diseases, and as a tool to combine recent advances in genome engineering of model organisms to uncover the biology of electric response.
Multi-phenotypic screening of multiple zebrafish larvae plays an important role in enhancing the quality and speed of biological assays. Many microfluidic platforms have been presented for zebrafish ...phenotypic assays, but multi-organ screening of multiple larvae, from different needed orientations, in a single device that can enable rapid and large-sample testing is yet to be achieved. Here, we propose a multi-phenotypic quadruple-fish microfluidic chip for simultaneous monitoring of heart activity and fin movement of 5-7-day postfertilization zebrafish larvae trapped in the chip. In each experiment, fin movements of four larvae were quantified in the dorsal view in terms of fin beat frequency (FBF). Positioning of four optical prisms next to the traps provided the lateral views of the four larvae and enabled heart rate (HR) monitoring. The device's functionality in chemical testing was validated by assessing the impacts of ethanol on heart and fin activities. Larvae treated with 3% ethanol displayed a significant drop of 13.2 and 35.8% in HR and FBF, respectively. Subsequent tests with cadmium chloride highlighted the novel application of our device for screening the effect of heavy metals on cardiac and respiratory function at the same time. Exposure to 5 $\mu$g/l cadmium chloride revealed a significant increase of 8.2% and 39.2% in HR and FBF, respectively. The device can be employed to monitor multi-phenotypic behavioral responses of zebrafish larvae induced by chemical stimuli in various chemical screening assays, in applications such as ecotoxicology and drug discovery.
In this paper, the heartbeat parameters of small model organisms, i.e. Drosophila melanogaster (fruit fly) and Danio rerio (zebrafish), were quantified in-vivo in intact larvae using microfluidics ...and a novel MATLAB-based software. Among different developmental stages of flies and zebrafish, the larval stage is privileged due to biological maturity, optical accessibility, and the myogenic nature of the heart. Conventional methods for parametric quantification of heart activities are complex and mostly done on dissected, irreversibly immobilized, or anesthetized larvae. Microfluidics has helped with reversible immobilization without the need for anesthesia, but heart monitoring is still done manually due to challenges associated with the movement of floating organs and cardiac interruptions. In our MATLAB software applied to videos recorded in microfluidic-based whole-organism assays, we have used image segmentation to automatically detect the heart and extract the heartbeat signal based on pixel intensity variations of the most contractile region of the heart tube. The smoothness priors approach (SPA) was applied to remove the undesired low-frequency noises caused by environmental light changes or heart movement. Heart rate and arrhythmicity were automatically measured from the detrended heartbeat signal while other parameters including end-diastolic and end-systolic diameters, shortening distance, shortening time, fractional shortening, and shortening velocity were quantified for the first time in intact larvae, using M-mode images under bright field microscopy. The software was able to detect more than 94% of the heartbeats and the cardiac arrests in intact Drosophila larvae. Our user-friendly software enables in-vivo quantification of D. melanogaster and D. rerio larval heart functions in microfluidic devices, with the potential to be applied to other biological models and used for automatic screening of drugs and alleles that affect their heart.
•An open-access software for rapid in-vivo quantification of larval heart function.•Microfluidics enabled desired orientation and reversible immobilization of larvae.•Image segmentation to automatically detect the heart and extract heartbeat signals.•Automatic heartbeat detection with more than 94% accuracy in intact larvae.•Semi-automatic measurement of heartrate and other heartbeat parameters.
Electrical stimulation of brain or muscle activities has gained attention for studying the molecular and cellular mechanisms involved in electric‐induced responses. We recently showed zebrafish's ...response to electricity. Here, we hypothesized that this response is affected by the dopaminergic signaling pathways. The effects of multiple dopamine agonists and antagonists on the electric response of 6 days‐postfertilization zebrafish larvae were investigated using a microfluidic device with enhanced control of experimentation and throughput. All dopamine antagonists decreased locomotor activities, while dopamine agonists did not induce similar behaviors. The D2‐selective dopamine agonist quinpirole enhanced the movement. Exposure to nonselective and D1‐selective dopamine agonists apomorphine and SKF‐81297 caused no significant change in the electric response. Exposing larvae that were pretreated with nonselective and D2‐selective dopamine antagonists butaclamol and haloperidol to apomorphine and quinpirole, respectively, restored the electric locomotion. These results reveal a correlation between electric response and dopamine signaling pathway. Furthermore, they demonstrate that electric‐induced zebrafish larvae locomotion can be conditioned by modulating dopamine receptor functions. Our electrofluidic assay has profound application potential for fundamental electric‐induced response research and brain disorder studies especially those related to the dopamine imbalance and as a chemical screening method when investigating biological pathways and behaviors.
Graphical and Lay Summary
The current work shows the first application of our multi‐fish microfluidic platform in chemical screening to answer novel questions such as examining if dopaminergic pathways are involved in zebrafish larvae electric response. Our chip can also be used by many in the fields of organism‐on‐a‐chip and zebrafish to investigate fundamental questions about zebrafish behavior or to perform larger‐scale neurobehavioral monitoring with quantitative phenotype readouts.
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