Microinjection Research Articles

Rice stripe virus (RSV) naturally infects rice plants through its insect vector during feeding. The small brown planthopper (SBPH) serves as the primary vector to transmit the viruses. The physical barriers preventing virus transmission in non-vectors have remained largely unexplored. In this study, we employed a hemocoel microinjection (MI) method to inoculate RSV into planthoppers and investigated viral infection progression in vector and non-vector species. Following microinjected into SBPHs, RSV successfully established infection across multiple tissues. Immunofluorescence analysis revealed progressive viral invasion, beginning with hemocytes and fat bodies, followed by salivary glands and gut tissues, achieving infection levels comparable to naturally viruliferous insects by 8 days. RSV-MI SBPHs demonstrated both horizontal transmission to rice plants and vertical transmission to offspring, effectively mimicking natural transmission patterns. When microinjected into two non-vector planthopper species, the white-backed planthopper and brown planthopper, RSV replicated only in hemolymph and established limited infection in hemocytes and fat bodies. This replication and accumulation did not affect insect survival. However, RSV failed to accumulate efficiently in salivary glands, preventing subsequent horizontal transmission to plants. MI-inoculated RSV could infect planthoppers, with different infection levels in vector and non-vector planthoppers. By comparing RSV infection in vector and non-vector species, our work offers novel insights into study of vector-virus specificity mechanisms. © 2025 Society of Chemical Industry.

We studied effects of microinjection (MI) into ovarian burses (OB) of infiltering agent dimethyl sulfoxide (DMSO) on number of ova shed (NOS) and estrous cycle (EC) of adult female rats with fourth day regular estrous cycles (REC), and we compared these effects with a MI of 100µg sulpiride solution in DMSO. Between 13:00- 14:00h of different days of EC, CER groups received 20µL of pure DMSO into both OB; the sham groups received MI with 20µL distillated water (H2O) into each OB. All rats with MI performed and an intact cyclic rat group were autopsied in the morning of next estrous vaginal shown (EVS). Both DMSO and H2O MI does not modify the EC duration and NOS. Other groups of cyclic rats received a MI with 20µL sulpiride solution into each OB and were autopsied at next EVS. Just the sulpiride MI performed on diestrous-1 day delayed 24h the next EVS, but that don’t occur in other sulpiride groups. The NOS were not modified in all experimental group. The DMSO direct administration in ovarian tissue does not affect gonadal primordial functions and its use is recommended like an infiltrating agent of non-polar drugs.

Gene-engineered animals created using gene-targeting technology have long been recognized as beneficial, valid, and valuable tools for exploring the function of a gene of interest, at least in early 2013. This approach, however, suffers from laborious and time-consuming tasks, such as the production of successfully targeted embryonic stem (ES) cells, their characterization, production of chimeric blastocysts carrying these gene-modified ES cells, and transplantation of those manipulated blastocysts to the recipient (pseudopregnant) females to deliver chimeric mice. Since the appearance of genome editing technology, which is now exemplified by the CRISPR/<em>Cas9</em> system, in late 2013, significant advances have been made in the generation of genome-edited animals through pronuclear microinjection (MI) of genome-editing components into fertilized eggs (zygotes) or electroporation (EP) of zygotes in the presence of these reagents. However, these procedures require the transfer of genome-edited embryos into the reproductive tracts of recipient females for further development. <u>G</u>enome editing via <u>o</u>viductal <u>n</u>ucleic <u>a</u>cids <u>d</u>elivery (GONAD) and its modified version, called “improved GONAD (<em>i</em>-GONAD),” were developed as an alternative to the MI- or EP-based genome-edited animal production and now recognized to be very convenient and straightforward as genome editing can only be performed <em>in</em> <em>vivo</em> (within the oviductal lumen where fertilized embryos exist). This system also enables the simultaneous transfection of epithelial cells <em>lining the oviductal lumen</em>. In this review, we summarize the recent advances in GONAD/<em>i</em>-GONAD and their derivatives and discuss the potential of these technologies to study various biological systems related to female reproduction.

The clustered regularly interspaced short palindromic repeats (CRISPR) technology has made it possible to produce genome-edited (GE) animals more easily and rapidly than before. In most cases, GE mice are produced by microinjection (MI) or by in vitro electroporation (EP) of CRISPR reagents into fertilized eggs (zygotes). Both of these approaches require ex vivo handling of isolated embryos and their subsequent transfer into another set of mice (called recipient or pseudopregnant mice). Such experiments are performed by highly skilled technicians (especially for MI). We recently developed a novelgenome editing method, called "GONAD (Genome-editing via Oviductal Nucleic Acids Delivery)," which can completely eliminate the ex vivo handling of embryos. We also made improvements to the GONAD method, termed "improved-GONAD (i-GONAD)." The i-GONAD method involves injection of CRISPR reagents into the oviduct of an anesthetized pregnant female using a mouthpiece-controlled glass micropipette under a dissecting microscope, followed by EP of the entire oviduct allowing the CRISPR reagents to enter into the zygotes present inside the oviduct, in situ. After the i-GONAD procedure, the mouse recovered from anesthesia is allowed to continue the pregnancy to full term to deliver its pups. The i-GONAD method does not require pseudopregnant female animals for embryo transfer, unlike the methods relying on ex vivo handling of zygotes. Therefore, the i-GONAD method can reduce the number of animals used, compared to the traditional methods. In this chapter, we describe some newer technical tips about the i-GONAD method. Additionally, even though the detailed protocols of GONAD and i-GONAD have been published elsewhere (Gurumurthy et al., Curr Protoc Hum Genet 88:15.8.1-15.8.12, 2016 Nat Protoc 14:2452-2482, 2019), we provide all the protocol steps of i-GONAD in this chapter so that the reader can find most of the information, needed for performing i-GONAD experiments, in one place.

Mosaicism - the presence of both wild-type and mutant alleles - is a serious problem for zygotic gene modification through gene editing using the Clustered regularly interspaced short palindromic repeats-Cas9 (CRISPR/Cas9) system. Different delivery methods, such as microinjection (MI), electroporation (EP), and transfection (TF), can be used to transfer CRISPR/Cas9 components into porcine zygotes. This study aimed to develop a method that combines MI, EP, and TF to improve mutation efficiency mediated through the CRISPR/Cas9 system for a triple-gene knockout in pigs. The study consisted of three groups: The MI group with three simultaneously microinjected guide RNAs (gRNAs) targeting α-1,3-galactosyltransferase (GGTA1), cytidine 32 monophosphate-N-acetylneuraminic acid hydroxylase (CMAH), and β-1,4-N-acetyl-galactosaminyltransferase 2 (B4GALNT2); the MI + EP group with two gRNAs targeting GGTA1 and B4GALNT2 genes delivered into zygotes through MI, followed by EP of gRNA targeting the CMAH 1 h later; and the MI + EP + TF group with MI of gRNA targeting GGTA1 gene into zygotes, followed by EP of gRNA targeting CMAH 1 h later, and then TF of gRNA targeting the B4GALNT2 gene into zona-free zygotes after another hour. The rate of blastocysts carrying mutations in one or two gene(s) was significantly higher in the MI + EP + TF group than in the MI group. However, the blastocyst formation rate of zygotes in the MI + EP + TF group was lower than that of the zygotes in the other treatment groups. The combination of CRISPR/Cas9 delivery methods may improve the mutation efficiency of triple-gene edited porcine blastocysts.

The rat is an important animal model for understanding gene function and developing human disease models. Knocking out a gene function in rats was difficult until recently, when a series of genome editing (GE) technologies, including zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the type II bacterial clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated Cas9 (CRISPR/Cas9) systems were successfully applied for gene modification (as exemplified by gene-specific knockout and knock-in) in the endogenous target genes of various organisms including rats. Owing to its simple application for gene modification and its ease of use, the CRISPR/Cas9 system is now commonly used worldwide. The most important aspect of this process is the selection of the method used to deliver GE components to rat embryos. In earlier stages, the microinjection (MI) of GE components into the cytoplasm and/or nuclei of a zygote was frequently employed. However, this method is associated with the use of an expensive manipulator system, the skills required to operate it, and the egg transfer (ET) of MI-treated embryos to recipient females for further development. In vitro electroporation (EP) of zygotes is next recognized as a simple and rapid method to introduce GE components to produce GE animals. Furthermore, in vitro transduction of rat embryos with adeno-associated viruses is potentially effective for obtaining GE rats. However, these two approaches also require ET. The use of gene-engineered embryonic stem cells or spermatogonial stem cells appears to be of interest to obtain GE rats; however, the procedure itself is difficult and laborious. Genome-editing via oviductal nucleic acids delivery (GONAD) (or improved GONAD (i-GONAD)) is a novel method allowing for the in situ production of GE zygotes existing within the oviductal lumen. This can be performed by the simple intraoviductal injection of GE components and subsequent in vivo EP toward the injected oviducts and does not require ET. In this review, we describe the development of various approaches for producing GE rats together with an assessment of their technical advantages and limitations, and present new GE-related technologies and current achievements using those rats in relation to human diseases.

Background and Aim:We previously developed the gene-editing by electroporation (EP) of Cas9 protein method, in which the CRISPR/Cas9 system was introduced into porcine in vitro fertilized (IVF) zygotes through EP to disrupt a target gene. This method should be further developed, and a combination of EP and MI methods should be evaluated in pigs. This study aimed to determine that a combination of microinjection (MI) and EP of CRISPR/Cas9 system could increase the rates of biallelic mutation for triple-gene knockout in porcine blastocysts. We targeted the pancreatic and duodenal homeobox1 (PDX1) gene using cytoplasmic MI 1 h before or after EP, which was used to edit alpha-1,3-galactosyltransferase (GGTA1) and cytidine 32 monophosphate-N-acetylneuraminic acid hydroxylase (CMAH) genes in porcine zygotes.Materials and Methods:We introduced guide RNAs targeting PDX1, GGTA1, and CMAH with the Cas9 protein into IVF zygotes (one-cell stage) through EP 10 h after the start of IVF (IVF; EP group) or in combination with MI (1 h before, MI-EP group, or after EP treatment EP-MI group) and evaluated the blastocyst formation rate and efficiency of target mutations in the resulting blastocysts.Results:Our results revealed a significant reduction in the rate of blastocyst formation in the two groups that underwent MI before and after EP (MI-EP and EP-MI group), compared with that in the groups treated with EP alone (EP group) (p=0.0224 and p<0.0001, respectively) and control (p=0.0029 and p<0.0001, respectively). There was no significant difference in the total mutation rates among the treatment groups in the resulting blastocysts. As an only positive effect of additional MI treatment, the rate of blastocysts carrying biallelic mutations in at least one target gene was higher in the MI-EP group than in the EP group. However, there was no difference in the rates of embryos carrying biallelic mutations in more than 2 target genes.Conclusion:These results indicate that although a combination of MI and EP does not improve the mutation efficiency or biallelic mutation for triple-gene knockout, MI treatment before EP is better to reduce mortality in porcine zygotic gene editing through a combination of MI and EP.

Induction of parturition in a large number of pregnant dairy goats and its benefits as a management tool in a commercial scale goat operation

The mechanisms underlying the establishment of left-right (L-R) asymmetry in bilaterians is one of the central enigmas in developmental biology. Amphioxus is an important model in studying the mechanisms of animal asymmetry specification due to its particular phylogenetic position, vertebrate-like embryogenesis and body plan. Recently, with the establishments of artificial breeding technology, high-efficiency microinjection method and gene knockout technology, researchers have successfully dissected the mechanisms of amphioxus L-R asymmetry development. In this review, we summarize the major progress in understanding L-R asymmetry specification in amphioxus and propose a model of regulation of L-R asymmetry in this species. Hh protein is transported dominantly to the right side by cilia movement, leading to R>L Hh signaling andCerexpression. Cer inhibits expression of Nodal, leading to the asymmetric expression of Nodal-dependent genes. The L-R differences in the propagation of the Nodal pathway result in the correct morphological L-R asymmetry development in amphioxus embryo. BMP signaling probably does not provide the asymmetric cue, but is necessary for correct expression ofCer andNodal.

Objective To compare the clinical effects of urokinase thrombolytic therapy for optic artery occlusion (OAO) and retinal artery occlusion (RAO) caused by facial microinjection with hyaluronic acid and spontaneous RAO. Methods From January 2014 to February 2018, 22 eyes of 22 patients with OAO and RAO caused by facial microinjection of hyaluronic acid who received treatment in Xi'an Fourth Hospital were enrolled in this retrospective study (hyaluronic acid group). Twenty-two eyes of 22 patients with spontaneous RAO were selected as the control group. The BCVA examination was performed using the international standard visual acuity chart, which was converted into logMAR visual acuity. FFA was used to measure arm-retinal circulation time (A-Rct) and filling time of retinal artery and its branches (FT). Meanwhile, MRI examination was performed. There were significant differences in age and FT between the two groups (t=14.840, 3.263; P=0.000, 0.003). The differecens of logMAR visual acuity, onset time and A-Rct were not statistically significant between the two groups (t=0.461, 0.107, 1.101; P=0.647, 0.915, 0.277). All patients underwent urokinase thrombolysis after exclusion of thrombolytic therapy. Among the patients in the hyaluronic acid group and control group, there were 6 patients of retrograde ophthalmic thrombolysis via the superior pulchlear artery, 6 patients of retrograde ophthalmic thrombolysis via the internal carotid artery, and 10 patients of intravenous thrombolysis. FFA was reviewed 24 h after treatment, and A-Rct and FT were recorded. Visual acuity was reviewed 30 days after treatment. The occurrence of adverse reactions during and after treatment were observed. The changes of logMAR visual acuity, A-Rct and FT before and after treatment were compared between the two groups using ttest. Results At 24 h after treatment, the A-Rct and FT of the hyaluronic acid group were 21.05±3.42 s and 5.05±2.52 s, which were significantly shorter than before treatment (t=4.569, 2.730; P=0.000, 0.000); the A-Rct and FT in the control group were 19.55±4.14 s and 2.55±0.91 s, which were significantly shorter than before treatment (t=4.114, 7.601; P=0.000, 0.000). There was no significant difference in A-Rct between the two groups at 24 h after treatment (t=1.311, P=0.197). The FT difference was statistically significant between the two groups at 24 h after treatment (t=4.382, P=0.000). There was no significant difference in the shortening time of A-Rct and FT between the two groups (t=0.330, 0.510; P=0.743, 0.613). At 30 days after treatment, the logMAR visual acuity in the hyaluronic acid group and the control group were 0.62±0.32 and 0.43±0.17, which were significantly higher than those before treatment (t=2.289, 5.169; P=0.029, 0.000). The difference of logMAR visual acuity between the two groups after treatment was statistically significant (t=2.872, P=0.008). The difference in logMAR visual acuity before and after treatment between the two groups was statistically significant (t=2.239, P=0.025). No ocular or systemic adverse reactions occurred during or after treatment in all patients. Conclusions Urokinase thrombolytic therapy for OAO and RAO caused by facial microinjection with hyaluronic acid and spontaneous RAO is safe and effective, with shortening A-Rct, FT and improving visual acuity. However, the improvement of visual acuity after treatment of OAO and RAO caused by facial microinjection with hyaluronic acid is worse than that of spontaneous RAO. Key words: Retinal artery occlusion/drug therapy; Thrombolytic therapy; Hyaluronic acid; Ophthalmic artery obstruction; Facial injections

We evaluated the function of dopaminergic receptors (DAR) of the anterior hypothalamus (AH) on the estral cycle (EC) regulation and spontaneous ovulation by a single microinjection (MI) with the dopaminergic antagonist haloperidol (HLP) in adult rats. One hundred thirty nine rats that exhibit forth-day estral cycles (cyclic animals: CA) received a stereotaxic surgery (STXS) on the right, left or both AH sides and were distributed in three different groups with a MI of 1 µL of: HLP (15 µg) or dimethyl-sulfoxide (vehicle) or other false MI group. All the animals with STXS were sacrificed in next vaginal estrus (VE) exhibited and the ova shed (OS) counted. In sixteen AC, the OS were counted at VE and forming a control group. The STXS affected the animals EC: just 59/139 exhibited a short EC (SEC) with 4.6±0.1 days compared with 80/139 that exhibited a long EC (LEC) of 13.6±0.2 days. False or HLP MI diminished OS just in animals exhibiting a SEC. STXS affects neuroendocrine processes controlling EC length when cutting dorsal connections to AH. The DAR of the AH participate on ovarian mechanisms of follicular selection.

Objective To investigate the efficacy of micro botulinum toxin A (BTX) combined with intense pulsed light (IPL) treatment in facial rejuvenation patients. Methods A total of thirty patients (1 male, 29 females, aged 35 to 55 years with average age of 45.56 years) with facial photoaging in the Department of Medical Cosmetology, Renmin Hospital of Wuhan University from Jan. to Sept. 2018 were enrolled as out-patients. Thirty patients were recruited in the study and randomly assigned averagely to the combination group and control group. All received standard IPL treatment every month for a total of 3 sessions. Patients in combined group received microinjections of diluted BTX on cheeks after the first IPL. VISIA CR images were analyzed before, 1 month and 3 months after treatment. Dermatologists assessed the improvement of color spots, wrinkles, texture, pore, and erythema improvement. The patient satisfaction survey was analyzed. Results The effective rate of the combined group was better than that of the intense pulsed light group one month after treatment (χ2=6.208, P=0.045), but there was no significant difference between the two groups three months after treatmen (χ2=1.077, P=0.584t). The patient satisfaction survey showed that there was no statistical difference between the two groups. Slight erythema and bruising were observed but disappeared at the end of the treatment (P>0.05). Conclusions The adjunctive use of MicroBTX can enhance treatment efficacy of IPL alone. It is a highly effective and safe treatment method for facial antiaging. Key words: Botulinum toxins, Type A; Injections; Micro injection; Intense pulsed light; Facial rejuvenation

Objective A practical length regulator of extended tube for microinjection pump was made and its clinical effect was observed. The method and basis were provided for the orderly management of various infusion pipelines in clinic. Methods The texture, inner diameter and color of the regulator are designed. Sixty patients who needed to use three or more lengthened tubes at the same time and were pumped through the same vein passage were randomly divided into two groups: the regulator group and the non-regulator group. 31 patients in the regulator group and 29 patients in the routine group were observed continuously for 3 days. During the study period, the number of tube shedding and the number of nursing errors were recorded. The average time required for the same dose of drugs to be pumped into the body was 10 ml/h, 25 ml/h and 50 ml/h, respectively. The average time spent by the nursing staff when the drugs were randomly changed or suspended was investigated in 52 cases. Staff satisfaction with its clinical application. Results The number of errors and tube shedding in the regulator group were 5 and 2 times respectively, and 34 and 9 times respectively in the routine group (P 0.05). The average time needed for drug replacement or suspension was 47.05 seconds in the regulator group and 91.68 seconds in the routine group (P < 0.05). The satisfaction of the two groups was 84.98 and 94.5 respectively (P < 0.05). Conclusion Compared with the conventional group, the regulator group can reduce the workload of nurses without increasing the risk of clinical use. The regulator is a safe, reliable and convenient pump pipe management device for pumps. Key words: Infusion pump; Systems; Extension tube; Regulator; Design; Utilization

Optogenetics is one of the biggest breakthroughs in neurobiology in recent decades, which is a revolutionary approach to precise intervention of specific cell functions through the combination of optical and genetic engineering techniques. However, this technique involves the interdisciplinary theoretical knowledge and methods, and it requires expensive equipments and long experimental period. Thus, it is not convenient to carry out widely in experiment teaching for undergraduates except the virtual simulation teaching. In present virtual simulation experiment, we aim to learn the basic principle, operation and application of optogenetics and achieve the desired teaching effects by simulating the process of the virus package, simulating the process of the animal selection, surgery and virus stereotactic microinjection, simulating the expression process of photosensitive channel, the extracellular verification process of photosensitive channel function in vivo, and simulating the observation process that optogenetic inhibition of glutamate neurons in the amygdala affect animal fear behavior. In this paper, the content design of the script making of the virtual experiment has been discussed in the above ways. Key words: Virtual simulation experiment; Experimental teaching; Optogenetics; Script

Objective To investigate the effects of orexin-A on firing activity of gastric distension-sensitive (GD) neurons in the basomedial amygdala (BMA) and food intake in diet-indaced obese rats. Methods Healthy male Wistar rats were selected, and the diet-induced obesity (DIO) rat model and diet-induced resistant (DR) rat model were established by high-fat diet. The effects of orexin-A and an opioid receptor antagonist naloxone on BMA GD neurons were observed by recording the extracellular potentials of single neurons.The effects of orexin-A and naloxone on the food intake of different rats were observed by using BMA catheterization.The mRNA expression and protein expression of orexin-1 receptor (OX-1R) and μ opioid receptor were detected by real-time PCR and Elisa, respectively. Results After microinjection of orexin-A into the BMA, the firing frequency of GD-sensitive neurons in the normal rats was significantly increased (GD-E: (78.3±6.9)%, GD-I: (55.5±4.7)%, P<0.01), and this effect was completely blocked by OX-1R receptor antagonist SB334867, and naloxone partially blocked the discharge-promoting effect of orexin-A; Compared with the normal rats, the firing frequency of GD-sensitive neurons in the DIO(GD-E: (91.6±7.1)%, GD-I: (67.9±8.1)%) and DR(GD-E: (87.9±6.8)%, GD-I: (69.2±5.8)%)rats was significantly increased after BMA injection of orexin-A (P<0.05). After administration of orexin-A into the BMA, food intake of the normal rats, DIO rats and DR rats ((2.38±0.34) g, (3.75±0.32) g, (4.01±0.38) g, respectively) was significantly increased (P<0.01), and the food intake of DR and DIO rats were significantly higher than that of normal rats (P<0.05). After BMA was injected with naloxone, the food intake of rats was inhibited, and the food intake of the DIO rats was significantly lower than that of the DR rats (P<0.05), food intake of the DR rats was significantly lower than that of the normal rats (P<0.05). The results of real-time PCR showed that the mRNA levels of OX-1R in DIO and DR rats were( 5.85±0.45 )and (6.03±0.42) were higher than that of normal rats, and the difference was significant (P<0.05); and mRNA levels of μ-opioid receptors in DIO and DR rats((4.51±0.42) and (8.31±0.41)times) were higher than those in normal rats (P<0.05). The results of Elisa showed that the protein levels of OX-1R in DIO ((2.98±0.28) ng/μl) and DR rats ((3.05±0.31) ng/μl) were higher than those in normal rats ((1.53±0.31)ng/μl, P<0.05). The content of μ-opioid receptor protein in DR rats ((4.21±0.35)ng/μl) was higher than that of DIO rats ((2.77±0.27) ng/μl), and higher than that of normal rats((1.48±0.32) ng/μl), the difference was significant (P<0.05). Conclusion BMA orexin-A promotes the spontaneous discharge of GD-sensitive neurons and food intake in normal rats, DIO rats and DR rats, μ-opioid receptors may be involved in the regulation of this process. Key words: Orexin-A; Food intake; Gastric distension-sensitive neurons; Diet-induced obesity rat; Amygdala

Lithium, a glycogen synthase kinase-3β (GSK-3β) inhibitor, prevents cannabinoid withdrawal syndrome, but there is limited data exploring the interaction between lithium and cannabinoid system on memory processes. The present study aimed to test the interaction between dorsal hippocampal (CA1 region) cannabinoid system and lithium on spatial memory in rats. Spatial memory was assessed in Morris Water Maze (MWM) apparatus by a single training session of eight trials. The results showed that pre-training intra-CA1 microinjection of ACPA, the cannabinoid type 1 receptor (CB1r) agonist, at doses of 0.001, 0.01 or 1 µg/rat, or AM251, the cannabinoid type 1 receptor (CB1r) antagonist, at doses of 1, 10 or 100 ng/rat, increased escape latency and traveled distance to the platform, suggesting a spatial learning impairment, whereas intraperitoneal administration of lithium (0.5, 1 or 5 mg/kg) had no effect on spatial learning. Also, rats that received lithium plus a lower dose of ACPA (0.001 µg/rat) or AM251 (1 ng/rat) had successful performance in the MWM. In the probe test, the results showed that pre-training administration of lithium (5 mg/kg) and ACPA (0.01 or 1 µg/rat) but not AM251 (at all doses used) impaired spatial memory retrieval. Also, lower dose of ACPA (0.001 µg/rat) or AM251 (1 ng/rat) potentiated the effect of ineffective doses of lithium (0.5 and 1 mg/kg) on spatial memory retrieval, while restored the effect of effective dose of lithium (5 mg/kg). In conclusion, cannabinoids may have a dual effect on lithium-induced spatial memory impairment in rats.

Objective To investigate the influences of anterior thalamic nucleus (ANT) stimulation on neurogenesis in hippocampus of epileptic rats. Methods Thirty-two male SD rats were randomly allocated to normal-control group(n=8), control-stimulation group(n=8), sham-stimulation group(n=8) and model-stimulation group(n=8). Eqileptic SD rat model was established by using microinjection of kainic acid in hippocampal CA3 area, and administered 48 h continuous ANT stimulation in the chronic stage.Epileptic seizures were monitored and counted.The levels of Ki-67, a neurogenesis protein in hippocampus was determined by Western blot and quantitative real-time polymerase chain reaction (qRT-PCR). Results The epilepsy seizure rate was (5.9±2.2) per week in the sham-stimulation group and (2.9±1.1) per week in model-stimulation group.Compared with sham-stimulated rats, ANT stimulation reduced seizures by 50.8% (P<0.05). Western blot analysis revealed that the relative levels of Ki-67 in the hippocampus of model-stimulation group significantly higher than that of the sham-stimulation group((0.44±0.15) vs (0.19±0.73), P<0.05). qRT-PCR analysis showed that relative levels of Ki-67 mRNA in the hippocampus of model-stimulation group were significantly higher than that of the sham-stimulation group((0.45±0.10) vs (0.15±0.06), P<0.05). Conclusion Chronic ANT stimulation can promote neurogenesis in epileptic rats, which may be a principle mechanism of the beneficial effect of ANT stimulation on epilepsy. Key words: Anterior thalamic nucleus; Electrical stimulation; Neurogenesis; Epilepsy

In order to investigate the effects of viscosity on spray formation and utilization of pesticide, different concentrations of xanthan gum (XG) were added into water and 0.1% Silwet 408 aqueous solution. Droplet size, relative span (RS), fan angle, length of breakup and maximum retention (Rm) were measured with the LU120-02 nozzle spraying under the pressure of 0.3 MPa. The dynamic spreading of the different solutions on maize leaves was tested using a 5 μL micro-injector. The results showed: VMD, RS, length of breakup and Rm went up as the increasing of XG concentration in the range of 0-0.5% with the same solution, while the fan angle of nozzle and spreading area on maize leaf showed the opposite tendency. Silwet 408 could reduce the surface tension of liquid, which could alter the dominant mode of spray formation and lead to earlier sheet breakup, especially in low viscosity solutions. Under the same concentration of XG the addition of Silwet 408 could reduce the RS of drop size spectrum but has no effect on VMD or fan angle. In water solution, there was no difference with different concentrations of XG in the spreading time on maize leaf. Besides, in the 0.1% Silwet 408 aqueous solution, the spraying time and area were several-fold of that in water with same XG concentration. Moreover, with the same XG concentration, the smaller surface tension liquid indicated lower Rm, and the difference was magnified as the concentration increases. This work has demonstrated that initial spray characteristics such as droplet size and RS, fan angle, length of breakup, Rm and spreading area can vary depending on the viscosity of spray liquids. Therefore, by transforming the viscosity of the spray liquid to adjust the droplet spectrum to reduce drift, increasing the Rm and spreading area to improve liquid utilization and reduce the usage of pesticides. Keywords: atomization, xanthan gum, spray, droplet size, breakup length, dynamic spreading DOI: 10.25165/j.ijabe.20181103.3802 Citation: Wang S L, He X K, Song J L, Wang S S, Jia X M, Ling Y. Effects of xanthan gum on atomization and deposition characteristics in water and Silwet 408 aqueous solution. Int J Agric & Biol Eng, 2018; 11(3): 29–34.

Objective To explore the better humidification oxygen therapy for patients with artificial airway from weaning to extubation, ensure the best humidification effect, keep airway unobstructed, shorten tubulization time and reduce the incidence of infection. Methods A total of 133 patients with artificial airway during weaning from ventilation admitted from March to December in 2016 in intensive care unit of the Second Affiliated Hospital of Chongqing Medical University were included in the study. They were divided into the experimental group (69 patients) and the control group (64 patients) by random lottery form. The experimental group was given improved combination device (venturi, heated humidifier and ventilator tube)during oxygen therapy for humidification and heating, while the control group was treated with oxygen therapy in endotracheal tube and continuous wet micro-injection pump 0.45% sodium chloride method. The heart rate, respiratory rate, blood oxygen saturation, offline time with tube, offline failure rate, sputum viscosity, sputum scab formation, irritant cough and pulmonary infection were compared between the two groups. Results The heart rate, respiratory rate, blood oxygen saturation and offline time with tube in the experimental group were (80.50±7.07) times/min, (17.38±1.92) times/min, 0.98±0.01, and (1.58±1.06) days, and which were (88.50±3.07) times/min, (21.38±1.51) times/min, 0.96±0.01 and (3.00±1.09) days in the control group. The differences were statistically significant (t = 2.268-4.782, P < 0.05 or 0.01). The offline failure (2 cases), sputum scab formation (3 cases), irritant cough (4 cases) and pulmonary infection(4 cases) were less than 8 cases, 12 cases, 20 cases,12 cases in control group. The differences were statistically significant (χ2=4.652-14.545, P < 0.05 or 0.01). The sputum viscosity of Ⅰ, Ⅱand Ⅲ were 5 cases, 52 cases and 12 cases in the experimental group, which were better than 13 cases, 11 cases and 40 cases in the control group. The difference was statistically significant (Z = 3.385, P < 0.01). Conclusions The improved oxygen therapy heated humidify strategy can not only achieve satisfactory humidification effect, but also improve the success rate of offline machines, shorten tubulization time, promote the comfort and tolerance of patients, and reduce the occurrence of infection. Key words: Airway management; Artificial airway; Patients during weaning from ventilation; Airway humidification; Oxygen inhalation therapy

Objective To explore the relationship between myeloid differentiation factor 88 (MyD88)/NF-κB signaling pathway and sevoflurane-induced cognitive deficit in aging rats. Methods Twenty months old male SD rats were randomly divided into 3 groups (n=15): control (C group), sevoflurane treatment (S group), and sevoflurane plus ST2825 treatment (IS group). The rats in S and IS groups were subjected to inhalation of 4% sevoflurane for 6 h, but the rats in C group inhaled air only. The rats in IS group received microinjection of ST2825 via lateral ventricle 10 min before sevoflurane exposure. The cognitive function was assessed with Morris water maze test and open field test. NF-κB activity was assessed with EMSA assay. The expression of TNF-α and IL-1β mRNA in the hippocampus were assessed with real-time PCR assay. The expressions of amyloid β(Aβ)42 were assessed with Western blot assay. Results Compared with C group, there were significant increases of escape latency period, the time the rats spent in the central square, activity of NF-κB, levels of TNF-α mRNA, IL-1β mRNA and Aβ42, but significant decreases of the number of grid crossing and the number of rearing in S group (P<0.05). Compared with S group, there were significant decreases of escape latency period, the time spent in the central square, activity of NF-κB, levels of TNF-α mRNA, IL-1β mRNA and Aβ42, but increases of the number of grid crossing and the number of rearing in IS group (P<0.05). Conclusions Sevoflurane-induced cognitive dysfunction in aging rats could be associated with activation of MyD88/NF-κB signaling pathway. Key words: Sevoflurane; Myeloid differentiation factor 88; Cognitive dysfunction