Dissection Procedure Research Articles

Surgical Management of Carcinoma Vulva-Case Series and Review of Literature.

The objective of this study was to study the risk factors, management protocols, and the outcome of vulvar cancer cases over a period of 2years in a tertiary care hospital. This is a case series of early-stage vulvar cancer in the Department of Surgical Oncology in BL Kapur Superspeciality Hospital from Jan 2016 to date. Five patients with histologically proven diagnosis of early-stage vulvar cancer were included. The mean age for the diagnosis of vulvar cancer was 58years and the peak incidence was seen in postmenopausal age group. All of the cases were squamous cell carcinomas in stage IB except one which was a basisquamous variant. All cases were treated primarily with surgery and vulvar flap reconstruction. Adjuvant therapy was not given in any case. Cases were followed from 6months to date, and no recurrence noted. The limitations of the study were rarity of disease and less number of cases. As all the cases in our study were in early stage of disease (stages I and II), surgical treatment in the form of modified radical vulvectomy with B/L inguinofemoral lymph node dissection and oncoplastic procedure was the treatment modality chosen for all the patients.

Parotidectomy in patients with head and neck cutaneous melanoma with cervical lymph node involvement.

Parotidectomy in melanoma of the coronal scalp and face with clinically involved cervical lymph node metastasis is based on predicted cervical lymphatic drainage described by O'Brien. In total, 40 parotidectomies with en bloc therapeutic neck dissection were retrospectively analyzed. Lymphatic spread of melanoma to the parotid lymph nodes was observed in 10 of 40 specimens (25%). Eight of the 10 parotid-positive patients developed a recurrence vs 17 of the 30 parotid-negative patients (P = 0.28). There were no differences in overall survival, melanoma-specific survival, and disease-free survival between the parotid-positive and parotid-negative patients. Although in this series no survival differences were found, parotidectomy still merits a sustained role in therapeutic neck dissection procedures to improve regional control and to prevent facial nerve damage after surgery for a second relapse from occult metastases in the parotid.

Real-Time Ex-Vivo Magnetic Resonance Image—Guided Dissection of Human Brain White Matter: A Proof-of-Principle Study

Modern neuroanatomic education should be based on interdisciplinary methods that allow an understanding of the cerebral circuitry, which is at the base of the structural connectivity. Ex-vivo MRI-guided dissection is an essential method for developing and refining the knowledge of complex 3-dimensional brain anatomy and the mutual relationships between structures and architecture of the white matter bundles. The aim of this technical note is to present a new and innovative method of studying human brain white matter. Four adult human cerebral hemispheres were prepared according to the Klinger's method. T1-weighted and T2-weighted and fluid attenuated inversion recovery images were obtained with a 3T magnetic resonance machine. The dissection was performed in a dedicated neurosurgical laboratory equipped with a microscope and an electromagnetic neuronavigation system that guided the whole white matter dissection. Gyri and sulci morphology were studied in detail. The relations between superficial and inner structures were observed before and after the dissection. Gray matter was carefully removed with blunt dissectors, and the U-fibers were exposed. Afterwards, deeper association and projection fibers, such as the arcuate fasciculus, superior and inferior longitudinal fasciculus, corona radiata, extreme and external capsule, claustrum, anterior commissure, and internal capsule were visualized under high magnification. The neuronavigation system was crucial for continuously checking the whole dissection procedure to avoid any accidental excision of fibers. Image-guided neuronavigated dissection can significantly improve the quality of white matter dissection and represents a valid tool for learning the 3-dimensional anatomy of the human brain tracts.

Quantitative Analysis of Neuronal Dendritic Arborization Complexity in Drosophila.

Dendrites are the branched projections of a neuron, and dendritic morphology reflects synaptic organization during the development of the nervous system. Drosophila larval neuronal dendritic arborization (da) is an ideal model for studying morphogenesis of neural dendrites and gene function in the development of nervous system. There are four classes of da neurons. Class IV is the most complex with a branching pattern that covers almost the entire area of the larval body wall. We have previously characterized the effect of silencing the Drosophila ortholog of SOX5 on class IV neuronal dendritic arborization complexity (NDAC) using four parameters: the length of dendrites, the surface area of dendrite coverage, the total number of branches, and the branching structure. This protocol presents the workflow of NDAC quantitative analysis, consisting of larval dissection, confocal microscopy, and image analysis procedures using ImageJ software. Further insight into da neuronal development and its underlying mechanisms will improve the understanding of neuronal function and provide clues about the fundamental causes of neurological and neurodevelopmental disorders.

Transmission Electron Microscopy of the Phloem with Minimal Artifacts.

It is a universal feature of seed plants that their phloem consists of a continuous sieve-tube system throughout the plant that is highly pressurized by its sugar contents. Cellular continuity and the pressure flow, osmotically generated in the source leaves, allow the assimilates to reach all sinks organs. However, both phloem features, the cellular continuity and the high pressure, are challenges when fixing the phloem for transmission electron microscopy. With very few exceptions, the tissue preparation necessary for the fixation evokes rapid wound responses that eventually result in artifacts.This chapter describes the steps necessary to minimize development of artifacts in the phloem and includes preparation of fixatives, a dissection procedure that optimizes penetration of the fixatives and application to axial and lateral plant organs. Moreover, as alternative to the established fixation of fresh hand sections, we suggest a xylem-assisted perfusion fixation method for herbaceous plants. After the initial fixation, the subsequent dehydration, embedding, and ultrathin sectioning of the material follow routine procedures, which are briefly discussed, as is the orientation of samples for obtaining transverse and longitudinal phloem sections.

Endoscopic ultrasonography-guided poly (lactic acid-co-glycolic acid)-poly (ethylene glycol)-poly (lactic acid-co-glycolic acid) thermogel tunnel creation for natural orifice transluminal endoscopic surgery in porcine model.

Natural orifice transluminal endoscopic surgery (NOTES) is a minimal invasive treatment. However, tissue dissection under endoscopy is still challenging due to the flexibility of endoscopy body and there is still no effective method for establishing a tunnel towards the targeted area. We previously showed that a new kind of thermogel could be submucosallly injected and served as a cushion for endoscopic dissection. Thus, in this study we investigated the feasibility and safety of tunnel creation using poly (lactic acid-co-glycolic acid)-poly (ethylene glycol)-poly (lactic acid-co-glycolic acid) (PLGA-PEG-PLGA) thermogel for NOTES in a porcine model. We prepared an injectable thermogel composed of PLGA-PEG-PLGA triblock copolymers which exhibited a low-viscous sol at room temperature and spontaneously transformed into a no-flowing gel at body temperature. This thermogel was used in NOTES in pigs. The success rate and adverse events were observed. The PLGA-PEG-PLGA thermogels were successfully injected to the targeted areas under the guide of endoscopic ultrasonography and the tunnels were created by sucking the gel during NOTES as the endoscopy went forwards in all the three animals. The necropsy of the pigs showed no evidence of iatrogenic injury. No serious bleeding and perforation was observed. The results demonstrated that thermogel injection and tunnel creation by suction during NOTES were feasible, which simplified the procedure of tissue dissection and developed a new method of identifying the targeted area for surgical interventions without causing severe tissue damage. The application of thermogel for tunnel creation in NOTES could optimize current procedures and may have a promising prospect in clinical application.

MUSCULAR ABNORMALITIES IN SUBMENTAL TRIANGLE – VARIATIONS OF ANTERIOR BELLY OF DIGASTRIC MUSCLE

The aim of this study is to highlight some unusual morphology of the digastric muscle anterior belly. During dissection of the bodies used in the practical lessons for second year medical students we found abnormal pattern of the suprahyoid region in the necks of two adult male cadavers. Both were with variable anterior bellies of digastric muscles. Case 1 presented a unilateral left variation, with an anomalous anterior belly. Case 2 presented bilateral variation, with the presence of two supernumerary bellies. Though multiplicity of anterior belly is common we provide interesting variations with anatomical and clinical importance. The different variants of the anterior belly of the digastric muscle can be found during the routine dissection or surgical procedure or imaging techniques. Anatomical variations of the anterior bellies of the digastric muscles can be easily confused with pathological conditions in CT and MR imaging. It is necessary to recognize that variants of the digastric muscle may happen in order to escape confusion when diagnosing abnormal lesions of the floor of the mouth and submental region.

Minimally invasive neck dissection: A 3-year retrospective experience of 45 cases.

Objective:Robot-assisted neck dissection requires a larger wound, is expensive and requires specialised equipment which is not easily available. We have developed an inexpensive minimally invasive neck dissection (MIND) procedure using simple endoscopic instruments in the past. This study was conducted to evaluate the safety, efficacy and reproducibility of the technique.Materials and Methods:From January 2013 to December 2016, we performed MIND on 45 patients with oral cancer using the standard endoscopic equipment. CO2 gas insufflation was used to create the working space. Intra-operative data, post-operative data and pathological characteristics were evaluated and overall survival (OS) and disease-free survival (DFS) Kaplan–Meier curves were compared using the Log-Rank test.Results:Median operative time was 130 (80–190) min with a mean blood loss of 63 (20–150) ml. Major intra-operative complications were not observed. The median number of nodes retrieved was 14 (range: 7–38). Three patients with a positive lymph node were advised to undergo adjuvant radiotherapy. After consultation, 12 out of 13 tongue cancer patients with a tumour depth >3 mm underwent adjuvant radiotherapy. Mean follow-up period was 31.5 (95% confidence interval [CI] 27.9–35.1) months and 27.8 (95% CI 23.6–32.1) months for OS and DFS, respectively. Four (8.9%) deaths and 8 (17.8%) recurrences were observed. The 3-year OS and DFS was 91.1% and 82.2%, respectively.Conclusion:MIND is aesthetically better than conventional procedures for oral cancer patients due to its safety, efficacy and reproducibility at any centre using the standard laparoscopic equipment.

A rare case of false positive metastatic nodes detected at PET/CT scan six months after the laparoscopic treatment of a patient with endometrial cancer

A 61-year-old woman was admitted to this hospital with a diagnosis of a poorly differentiated endometrioid carcinoma with myometrial invasion greater than 50 % (IBG3) at MRI. A laparoscopic surgical treatment with peritoneal washing, a type A radical hysterectomy, bilateral salpingo-oophorectomy, and pelvic and para-aortic lymphadenectomy were programmed. Six months later the patient underwent a PET/CT scan that diagnosed a suspected recurrence into the lymph node tissue laterally to the left common iliac artery. Then, the patient underwent a second laparoscopic procedure for dissection of the positive tissue at PET/CT scan on the external site of common iliac artery. At histological examination the dissected two lymph nodes were negative for metastasis and the patient was regularly discharged three days after laparoscopy.

Prospective multicenter study for robotic anatomical major liver resection : Can robotic beginners safely perform ?

Introduction: The robotic surgical system had been widely accepted in various surgical field with the expectations of overcoming the limitation of laparoscopic surgery. However, robotic liver resection had not generalized, so far. Thus, this study aimed to evaluate the feasibility and safety of robotic major liver resection by a prospective multicenter study. Method: From July 2017 to December 2018, five surgeons who were the novice in robotic liver resection but experienced a lot in open and laparoscopic liver resection in five tertiary hospitals performed 46 cases of robotic major anatomical liver resection. Perioperative patient’s clinical data and surgical data including detailed procedure time were prospectively collected. All operations were performed identical procedures for dissection and hemostasis and were a totally robotic approach. Result: Twenty-two cases of left hemihepatectomy, 1 extended left hemihepatectomy, 14 right hemihepatectomy, 2 right anterior sectionectomy, 6 right posterior sectionectomy, and one central bisectionectomy were performed. The most common indications were hepatocellular carcinoma for 21 cases following intrahepatic cholangiocarcinoma for 7 cases, liver metastases for 3 cases. Surgical resection margins for all tumor cases were negative. Total average operation time was 378.58 ± 124.31 (190∼696) minutes and estimated intraoperative blood loss was 276.67 ± 397.41 mL (range, minimal to 2600mL). Overall complications were developed in 16 cases (34.8%). In terms of severe (Clavien-Dindo classification, more than III) surgical complications, there were 3 cases of postoperative fluid collection treated with drainage and one case of bile leakage treated with percutaneous transhepatic biliary drainage. Only one case out of 46 cases was converted to the conventional open left hemihepatectomy because of bleeding. Transfusion was done in 2 patients (4.3%) and Pringle maneuver was applied in 16 (34.8%). Mean hospital stay was 7.3 days. Conclusion: Robotic anatomic major liver resection might be safely performed even by robotic beginners but experienced open liver surgeons.

Prolonged occlusion of the hepatoduodenal ligament to reduce risk of bleeding and tumor spread during recipient hepatectomy for living donor liver transplantation

Introduction: Prevention of excessive bleeding during liver transplantation (LT) operations presents a major challenge. Compared to deceased donor LT, living donor LT (LDLT) is more vulnerable to bleeding because of additional dissection procedures. We herein introduce our technique for prolonged occlusion of the hepatoduodenal ligament applied to recipient hepatectomy for LDLT. Method: Simulated assessment of splanchnic hemodynamics on prolonged occlusion of the hepatoduodenal ligament showed that patients with cirrhotic liver appeared to tolerate the procedure as like in the patients with normal liver. We accumulated experience on the prolonged Pringle maneuver with curved intestinal clamps. Result: This technique was applied to more than 60 cases of adult LDLT operations from early 2014 until the end of 2018. Initially, application of this technique was limited to patients showing heavy bleeding during perihilar mobilization. Thereafter, this technique was applied at the start of liver mobilization and stopped after complete mobilization of the retro-hepatic vena cava. Recently, this technique was also applied during dissection of the hepatoduodenal ligament. The mean total occlusion duration was 67±13 minutes. No patient suffered from major serosal peritoneal tearing-associated bleeding or hepatic artery problems. Intentional prolonged occlusion over two hours was applied to five patients who had advanced hepatocellular carcinoma. Conclusion: We believe that prolonged occlusion of the hepatoduodenal ligament is a simple effective method to reduce intraoperative bleeding, and that it has potential benefit to reduce the risk of intraoperative tumor spread during LDLT operations.

Prolonged occlusion of the hepatoduodenal ligament to reduce risk of bleeding and tumor spread during recipient hepatectomy for living donor liver transplantation.

Backgrounds/AimsPrevention of excessive bleeding during liver transplantation (LT) operations presents a major challenge. Compared to deceased donor LT, living donor LT (LDLT) is more vulnerable to bleeding because of additional dissection procedures. We herein introduce our technique for prolonged occlusion of the hepatoduodenal ligament applied to recipient hepatectomy for LDLT.MethodsSimulated assessment of splanchnic hemodynamics on prolonged occlusion of the hepatoduodenal ligament showed that patients with cirrhotic liver appeared to tolerate the procedure as like in the patients with normal liver. We accumulated experience on the prolonged Pringle maneuver with curved intestinal clamps.ResultsThis technique was applied to more than 60 cases of adult LDLT operations from early 2014 until the end of 2018. Initially, application of this technique was limited to patients showing heavy bleeding during perihilar mobilization. Thereafter, this technique was applied at the start of liver mobilization and stopped after complete mobilization of the retro-hepatic vena cava. Recently, this technique was also applied during dissection of the hepatoduodenal ligament. The mean total occlusion duration was 67±13 minutes. No patient suffered from major serosal peritoneal tearing-associated bleeding or hepatic artery problems. Intentional prolonged occlusion over two hours was applied to five patients who had advanced hepatocellular carcinoma.ConclusionsWe believe that prolonged occlusion of the hepatoduodenal ligament is a simple effective method to reduce intraoperative bleeding, and that it has potential benefit to reduce the risk of intraoperative tumor spread during LDLT operations.

Remote-Access Thyroidectomy: A Multi-Institutional North American Experience with Transaxillary, Robotic Facelift, and Transoral Endoscopic Vestibular Approaches

Many remote-access approaches (RAAs) to the thyroid have been described to circumvent anterior neck scarring, including the transaxillary, robotic facelift, and transoral endoscopic vestibular approaches. These techniques have been popularized in Asia, but adoption has been slow in North America. We aimed to examine multi-institutional North American outcomes with RAA thyroidectomy in the context of these institutions' transcervical approach (TCA) outcomes. Cases of lobectomy and total thyroidectomy via transaxillary, robotic facelift, and transoral endoscopic vestibular approaches were reviewed. Demographic characteristics, outcomes, and complications were compared with the same measures in patients undergoing lobectomy and total thyroidectomy via TCA by the primary RAA surgeons at each institution. Patients who underwent parathyroidectomy or other concomitant neck dissection procedures were excluded. Two hundred and sixteen RAA thyroidectomies were attempted (92 transoral endoscopic vestibular approaches, 70 transaxillary, and 54 robotic facelift) and 410 TCA thyroidectomies were performed. There was no difference in mean index nodule sizes between RAA (2.8 ± 1.6 cm) and TCA (2.9 ± 1.9 cm) cohorts (p= 0.72). Median operative times for lobectomy were 146 minutes (range 60 to 343 minutes) and 90 minutes (range 25 to 247 minutes) for the RAA and TCA cohorts, respectively (p < 0.0001). Median operative time for total thyroidectomy was 170 minutes (range 100 to 398 minutes) vs 126.5 minutes (range 51 to 260 minutes) for the RAA and TCA cohorts, respectively (p < 0.0001). There was no difference in the rates of permanent recurrent laryngeal nerve injury between the RAA (0 of 216 [0%]) and TCA (0 of 410 [0%]) cohorts (p= 0.99). Remote-access approach thyroidectomy can be performed in a select North American patient population with outcomes comparable with TCA.

Novel three-dimensional imaging system may facilitate gastric endoscopic submucosal dissection procedure: an ex vivo animal study.

Background and study aims Endoscopic submucosal dissection (ESD) requires advanced skills to perform safely without complications. The current study evaluated the usefulness of a novel three-dimensional (3D) imaging system in ESD using porcine stomachs. Methods Four endoscopists (two trainees and two experts) performed eight ESD procedures using both 3D and 2D images. The usefulness of 3D image versus 2D image was evaluated by visibility and procedure time. In addition, occurrence of eyestrain and dizziness in 3D image was assessed. Results En bloc resection was successfully achieved, without perforation, in all cases. The evaluation score in the 3D image group was better than that in the 2D image group, particularly depth perception was statistically significantly good. No significant difference was found in the working speed between the 2D and 3D image groups. Two examiners experienced eyestrain and dizziness while using the 3D image. Conclusions All the ESD procedures were performed safely. Depth perception using the 3D image was better than with the 2D image. A novel 3D image system may facilitate ESD.

Safe and effective use of free tissue transfer for facial reanimation in complex facial palsy

In irreversible, dense facial paralysis, facial reanimation surgery to recreate the smile is established as the gold standard. Most patients presenting for treatment will have had little or no prior surgical interventions. A significant minority may have undergone prior surgery, making any attempt at facial reanimation complex. These include patients with previous free flap and neck dissection procedures and where multiple prior procedures have been undertaken, including attempts at facial reanimation. Occasionally, a composite tissue reconstruction is required. In these complex cases, it is challenging to determine the best surgical solution for facial reanimation. Microsurgical techniques utilising free tissue transfer may seem to be relatively contraindicated. A series of 15 cases where free tissue transfer was used successfully for facial reanimation in complex cases is presented. These patients were otherwise deemed to be suitable for facial reanimation and were defined as complex utilising defined criteria. Results were assessed by using the Sunnybrook scoring system where possible and in all cases by measuring symmetry at rest and modiolar excursion when smiling. Standardised pre- and post-operative photos and videos were also used. The average improvement in Sunnybrook score was 39. The average improvement in resting symmetry was 8.5 mm. Thirteen patients developed facial movement corresponding to a good-quality smile with average improvement of modiolus movement of 6 mm. There were no free flap failures. These results demonstrate that even very complex cases of facial paralysis can be effectively and safely treated with free tissue transfer for facial reanimation. Level of Evidence: Level IV

Transanal endoscopic microsurgical submucosa dissection in the treatment of rectal adenomas and T1 rectal cancer.

BackgroundThe treatment of flat rectal adenomas is challenging. The technical difficulty and the potential of malignancy in suspected benign lesions are the factors in question. Surgical and interventional endoscopic techniques are implemented in Europe without a clear strategy. To minimize recurrent adenoma and unclear histopathological work up en bloc excision is desirable.Methods and resultsWe demonstrate in this article the transanal endoscopic microsurgical submucosa dissection (TEM-ESD) procedure as a feasible method for en bloc excision of rectal adenomas and early rectal cancer. The surgical technique is demonstrated in detail with the help of a video of the operation that is available online. The results of a consecutive series of 78 patients are presented.ConclusionTEM-ESD is a safe procedure for resection of rectal adenomas and low risk carcinomas. It offers the possibility of organ preservation and minimizes functional disturbances. In case of a necessary salvage operation, the preserved integrity of the rectal muscle tube grants maximal oncological safety.Electronic supplementary materialThe online version of this article (10.1007/s00053-018-0291-3) includes a video on the surgical technique: TEM ESD. The article and supplemental material are available at http://www.springermedizin.de/der-chirurg. The supplemental material can be found at the end of the article under “Supplementary material”.

PS02.002: EN-BLOC MEDIASTINAL LYMPH NODE DISSECTION USING A LAPAROSCOPIC TRANSHIATAL APPROACH FOR ESOPHAGEAL AND ESOPHAGOGASTRIC JUNCTION CANCERS

Abstract Background The procedure for mediastinal lymph node dissection using a laparoscopic transhiatal approach (LTHA) had not been established for esophageal and esophagogastric junction (EGJ) cancers because of the difficulties associated with surgery. We developed a novel and simple technique for their en-bloc dissection using LTHA. To date, 296 patients had undergone our method during various esophageal surgical procedures, including 39 cases with EGJ cancer. We describe our technique and evaluate the optimal range of mediastinal lymph node dissection for EGJ cancer. Methods The esophageal hiatus was opened and CO2 was introduced into the mediastinum. The posterior plane of the pericardium was extended, and the anterior side of the subcarinal, main bronchial, thoracic paraaortic and pulmonary ligament lymph nodes were separated. The posterior side of these lymph nodes was then separated. Finally, while lifting lymph nodes like a membrane, they were resected from bilateral mediastinal pleura, main bronchi and tracheal bifurcation. Results 1) Patients with EGJ cancers performed middle and lower mediastinal lymph node dissection by LTHA (n = 39) were compared with those performed by the right thoracotomy (n = 41). The total operative time and bleeding were significantly decreased by LTHA. The number of resected middle and lower mediastinal lymph nodes, pR0 rate, and mediastinal recurrence in the two groups were not different. In 87.2% of patients treated by LTHA, extubation was performed at 0 POD. Postoperative respiratory complications was decreased by LTHA (LTHA: 7.7%, thoracotomy: 17.0%). 2) Patients with EGJ cancers performed total mediastinal lymph node dissection by thoracotomy for the time before introducing LTHA were analyzed (n = 37). Upper and/or middle mediastinal lymph nodes metastasis was observed in 10 cases, and all of them had advanced tumors. All of 6 cases with upper mediastinal lymph nodes metastasis were squamous cell carcinoma. Even in adenocarcinoma, middle mediastinal lymph node metastasis was observed in 2 cases, suggesting the importance of mediastinal lymph node dissection for advanced EGJ cancers. Conclusion Our surgical procedure resulted in a good surgical view, safe en-bloc mediastinal lymph node dissection, and the decrease of postoperative respiratory complications. Disclosure All authors have declared no conflicts of interest.

Isolation of Mandibular Gland Reservoir Contents from Bornean 'Exploding Ants' (Formicidae) for Volatilome Analysis by GC-MS and MetaboliteDetector.

The aim of this manuscript is to present a protocol describing the metabolomic analysis of Bornean 'exploding ants' belonging to the Colobopsis cylindrica (COCY) group. For this purpose, the model species C. explodens is used. Ants belonging to the minor worker caste possess distinctive hypertrophied mandibular glands (MGs). In territorial combat, they use the viscous contents of their enlarged mandibular gland reservoirs (MGRs) to kill rival arthropods in characteristic suicidal 'explosions' by voluntary rupture of the gastral integument (autothysis). We show the dissection of worker ants of this species for the isolation of the gastral portion of the wax-like MGR contents as well as listing the necessary steps required for solvent-extraction of the therein contained volatile compounds with subsequent gas chromatography-mass spectrometry (GC-MS) analysis and putative identification of metabolites contained in the extract. The dissection procedure is performed under cooled conditions and without the use of any dissection buffer solution to minimize the changes in the chemical composition of the MGR contents. After solvent-based extraction of volatile metabolites contained therein, the necessary steps for analyzing the samples via liquid-injection-GC-MS are presented. Lastly, data processing and putative metabolite identification with the use of the open-source software MetaboliteDetector is shown. With this approach, the profiling and identification of volatile metabolites in MGRs of ants belonging to the COCY group via GC-MS and the MetaboliteDetector software become possible.

Isolation of Mandibular Gland Reservoir Contents from Bornean 'Exploding Ants' (&lt;em&gt;Formicidae&lt;/em&gt;) for Volatilome Analysis by GC-MS and MetaboliteDetector

The aim of this manuscript is to present a protocol describing the metabolomic analysis of Bornean 'exploding ants' belonging to the Colobopsis cylindrica (COCY) group. For this purpose, the model species C. explodens is used. Ants belonging to the minor worker caste possess distinctive hypertrophied mandibular glands (MGs). In territorial combat, they use the viscous contents of their enlarged mandibular gland reservoirs (MGRs) to kill rival arthropods in characteristic suicidal 'explosions' by voluntary rupture of the gastral integument (autothysis). We show the dissection of worker ants of this species for the isolation of the gastral portion of the wax-like MGR contents as well as listing the necessary steps required for solvent-extraction of the therein contained volatile compounds with subsequent gas chromatography-mass spectrometry (GC-MS) analysis and putative identification of metabolites contained in the extract. The dissection procedure is performed under cooled conditions and without the use of any dissection buffer solution to minimize the changes in the chemical composition of the MGR contents. After solvent-based extraction of volatile metabolites contained therein, the necessary steps for analyzing the samples via liquid-injection-GC-MS are presented. Lastly, data processing and putative metabolite identification with the use of the open-source software MetaboliteDetector is shown. With this approach, the profiling and identification of volatile metabolites in MGRs of ants belonging to the COCY group via GC-MS and the MetaboliteDetector software become possible.

Circadian rhythm bifurcation induces flexible phase resetting by reducing circadian amplitude.

Shift-work and jet-lag-related disorders are caused by the limited flexibility of the suprachiasmatic nucleus (SCN), a master circadian clock in the hypothalamus, to adjust to new light-dark (LD) cycles. Recent findings confirmed here establish that behavioral jet lag after simulated time-zone travel is virtually eliminated following bifurcated circadian entrainment under a novel and atypical 24-h light:dark:light:dark (LDLD) cycle. To investigate the mechanisms of this fast resetting, we examined the oscillatory stability of the SCN and peripheral tissues in LDLD-bifurcated mice employing the dissection procedure as a perturbing resetting stimulus. SCN, lung, liver, and adrenal tissue were extracted at times throughout the day from female and male PER2::Luciferase knock-in mice entrained to either LDLD or a normal LD cycle. Except for adrenals, the phase of the cultured explants was more strongly set by dissection under LDLD than under normal LD. Acute bioluminescence levels of SCN explants indicate that the rhythm amplitude of PER2 is reduced and phase is altered in LDLD. Real-time quantitative PCR suggests that amplitude and rhythmicity of canonical clock genes in the lung, liver, and kidney are also significantly reduced in LDLD in vivo. Furthermore, spatiotemporal patterns of PER2 peak time in cultured SCN were altered in LDLD. These results suggest that altered gene expression patterns in the SCN caused by bifurcation likely result in fast resetting of behavior and cultured explants, consistent with previously reported mathematical models. Thus, non-invasive, simple light manipulations can make circadian rhythms more adaptable to abrupt shifts in the environmental LD cycle.