The ATPase activity of the phosphatidylethanolamine flippase TAT-5 inhibits extracellular vesicle budding from the plasma membrane.

Cells release extracellular vesicles (EVs) from their surface, but the mechanisms that govern EV release by plasma membrane budding are poorly understood. The lipid flippase TAT-5 inhibits EV release from the plasma membrane in C. elegans , but how the level of flippase activity regulates EV release was unknown. We generated point mutations in the DGET motif of TAT-5 predicted to lead to a partial or complete loss of ATPase activity. We discovered that tat-5(E246Q) mutants were sterile, while tat-5(D244T) mutants produced embryos that arrested during development. Using degron-based reporters, we found that EV release was increased in tat-5(D244T) mutant embryos and that phagocytosis was also disrupted. These data suggest that a low level of flippase activity can promote fertility, while a higher level of flippase activity is required to inhibit EV release, allow phagocytosis, and carry out embryonic development.

Conserved N- and C-terminal motifs of PAD-1 are required to inhibit extracellular vesicle release.

Cells release extracellular vesicles (EVs) carrying cargos that can influence development and disease, but the mechanisms that govern EV release by plasma membrane budding are poorly understood. We previously showed that the Dopey protein PAD-1 inhibits EV release from the plasma membrane in C. elegans . However, PAD-1 is large, and the domains required to regulate EV release were unknown. Here, we reveal that the conserved N-terminal EWAD motif and C-terminal leucine zippers are required to inhibit EV release from the plasma membrane. Revealing a role for these domains is an important first step to identifying how EV release is regulated.

Paediatric pelvic fractures - an updated literature review.

BACKGROUND: Pelvic fractures in children are indicative of significant trauma. Patients will often have associated injuries - some of which require urgent intervention to prevent death and disability. Paediatric and adult pelvises respond to traumatic forces differently and distinct approaches are required for each population. Historically, pelvic fractures have been treated conservatively, but this trend is changing with a better understanding of the pelvis' inability to remodel significant deformity, as well as new techniques for operative fixation. METHODS: A comprehensive search of the literature was conducted for articles published between 2000 and 2020 on paediatric pelvic fractures using medical databases including PubMed, Embase and the Cochrane Library. RESULTS: We included 143 studies in our literature review and summarized the incidence, pathophysiology, assessment, management and complications associated with paediatric pelvic fractures. CONCLUSIONS: The rarity of paediatric pelvic fractures corresponds with a paucity of randomized clinical trials covering this topic. Trends such as the screening pelvic x-ray are derived from adult populations but are now questioned in children. Other aspects of assessment and management of these children warrant such levels of scrutiny.

Paediatric intestinal pseudo-obstruction: a scoping review.

Paediatric intestinal pseudo-obstruction (PIPO) encompasses a group of rare disorders in which patients present with the clinical features of bowel obstruction in the absence of mechanical occlusion. The management of PIPO presents a challenge as evidence remains limited on available medical and surgical therapy. Parenteral nutrition is often the mainstay of therapy. Long-term therapy may culminate in life-threatening complications including intestinal failure-related liver disease, central line thrombosis and sepsis. Intestinal transplantation remains the only definitive cure in PIPO but is a complex and resource-limited solution associated with its own morbidity and mortality. We conducted a scoping review to present a contemporary summary of the epidemiology, aetiology, pathophysiology, diagnosis, management and complications of PIPO.Conclusion: PIPO represents a rare disorder that is difficult to diagnose and challenging to treat, with significant morbitity and mortality. The only known cure is intestinal transplantation. What is Known: • Paediatric intestinal pseudo-obstruction is a rare, heterogeneous disorder that confers a high rate of morbidity and mortality • Complications of paediatric intestinal pseudo-obstruction include chronic pain, small intestine bacterial overgrowth and malrotation. Other complications can occur related to its management, such as line infections with parenteral nutrition or cardiac side effects of prokinetic medications What is New: • Progress in medical and surgical therapy in recent years has led to improved patient outcomes • Enteral autonomy has been reported in most patients at as early as 1 month post-transplantation.

Paediatric adhesive bowel obstruction: a systematic review.

Adhesions following abdominal surgery remain a common cause of bowel obstruction. The incidence is between 1 and 12.6% in children who have had previous abdominal surgery. While conservative management is usually trialled in all patients (including children) suspected of having ASBO, the majority will require surgical intervention. New materials such as Seprafilm® have been studied in the paediatric population, with promising results of its use in index abdominal surgeries to prevent the formation of adhesions. In this article, we conducted a systematic review to present an overview of the current knowledge on the incidence, aetiology, pathophysiology, clinical presentation, and management of ASBO.

Paediatric chemical burns: a clinical review.

Although they account for a small proportion of burns in paediatrics, injuries from chemicals can be just as devastating as other mechanisms of burn injury. At least 25,000 chemicals exist which can cause burns: in children, they are often caused by household chemicals via accidental exposure. The mechanism by which corrosive substances produce chemical burns highlights the importance of early and plentiful irrigation of the burn area, removal of contaminated clothes and careful clinical assessment. Surgical intervention is uncommon but often follows the principles for thermal burns. This article reviews the aetiology, incidence, clinical presentation, management, complications and prevention of chemical burns. What is Known • Chemical burns in paediatrics are often caused by accidental exposure to chemicals available at home • Differences in the pathophysiology of chemical burns reinforces the need for early irrigation What is New • New irrigation fluids show promise in adults and need further study in children • The nature of chemical cutaneous burns can make assessment of wound depth difficult. Laser Doppler Imaging (LDI) is an accurate technique that can be used clinically to determine burn depth in thermal burns and is an area of future interest in the assessment of chemical burns.

Histopathologic changes in cadaver eyes after MicroPulse and continuous wave transscleral cyclophotocoagulation.

OBJECTIVE: The purpose of this study was to compare the acute histological effects of MicroPulse transscleral cyclophotocoagulation (MPCPC) using the MicroPulse P3 Device and continuous wave transscleral cyclophotocoagulation (CWCPC) on the ciliary body and adjacent structures in human cadaver eyes. METHODS: Quadrants of 6 human cadaver eyes from 3 different donors were subjected to traditional CWCPC, slow burn CWCPC, MPCPC, or no treatment (internal control). Sutures were used to differentiate different treatment areas on each eye. Differential inking was applied after treatments to aid in microscopic correlation. All specimens were subject to standard histologic processing. Tissue sections were cut at 4 microns and stained with hematoxylin and eosin according to established protocols. Pathologic evaluation by light microscopy was confirmed by a senior pathologist blinded to treatment groups. RESULTS: In all 6 eyes, tissues treated with traditional and low burn CWCPC showed variable coagulative tissue damage to the ciliary body compared with untreated tissues. Minimal histologic changes were identified within the ciliary processes, although variable pigment clumping and streaming were noted within the pigmented ciliary epithelium. In contrast to CWCPC, MPCPC-treated tissues showed only minimal coagulative tissue damage to the ciliary body. Variable pigment clumping and streaming, however, were also noted in the pigmented ciliary epithelium in MPCPC-treated tissues. CONCLUSIONS: In human cadaver eyes, MPCPC treatment caused less tissue disruption to the ciliary body compared with traditional and low burn CWCPC treatments. MPCPC may be a less destructive and more selective method of cyclophotocoagulation when compared with traditional and low burn CWCPC.

Optimizing Quality of Life for Patients with Breast Cancer-Related Lymphedema: A Prospective Study Combining DIEP Flap Breast Reconstruction and Lymphedema Surgery.

BACKGROUND: Patients with breast cancer-related lymphedema can be treated with a simultaneous deep inferior epigastric perforator (DIEP) flap, vascularized inguinal lymph node transfer, and lymphovenous anastomosis for aesthetic breast reconstruction and lymphedema in one operation. METHODS: The authors performed a comparison of prospectively followed patients who underwent free flap breast reconstruction with vascularized inguinal lymph node transfer and anastomosis to a retrospective cohort of patients who underwent free flap breast reconstruction with vascularized inguinal lymph node transfer alone. RESULTS: Thirty-three patients underwent DIEP flap reconstruction with vascularized inguinal lymph node transfer and lymphovenous anastomosis, and 21 received a free flap with lymph node transfer alone. There were no significant differences in demographics, adjuvant chemotherapy, or radiation therapy. The average number of nodes removed was also equivalent (21.2 versus 21.4 nodes). Two anastomoses per patient, on average, were performed (range, one to four) in the combined cohort, and all patients (100 percent) reported a subjective improvement in symptoms, compared with 81.0 percent of patients undergoing only lymph node transfer (p = 0.019). Perometer measurements demonstrated a significant reduction between the groups at early time points [3 months, 40.7 percent versus 20.0 percent (p = 0.037); 6 months, 57.0 percent versus 44.5 percent (p = 0.043)]; however, the difference was not statistically significant at 12 months (60.4 percent versus 57.8 percent; p = 0.43). CONCLUSION: This is the first prospective study demonstrating the safety and efficacy of a combined DIEP flap with vascularized inguinal lymph node transfer and lymphovenous anastomosis, which may be superior to lymph node transfer alone. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, III.

Early results of micropulse transscleral cyclophotocoagulation for the treatment of glaucoma.

PURPOSE: To describe our clinical experience with the efficacy and safety of micropulse transscleral cyclophotocoagulation as a treatment for glaucoma. METHODS: In this retrospective case series, we reviewed the charts of 95 consecutive patients with various glaucoma subtypes who underwent micropulse transscleral cyclophotocoagulation. Patients were offered micropulse transscleral cyclophotocoagulation if they had perimetric glaucoma refractory to intraocular pressure-lowering topical medications and who were poor candidates for traditional filtering surgery. Eligible patients were treated with the Micropulse P3 device (IQ 810 Laser Systems; Iridex, Mountain View, CA, USA) at 2.0-2.5 W for a duration of 90 s per hemisphere at a 31.3% duty cycle. If a retreatment was needed, the power was increased to up to 3.0 W with other parameters remaining the same. Patients were considered successfully treated if their intraocular pressure was lowered by at least 20% compared to their baseline. The main outcome measure was post-operative intraocular pressure; secondary outcome measures included the number of adverse events and complications that occurred with treatment. RESULTS: The glaucoma subtypes treated included primary open-angle glaucoma (n = 51), exfoliation glaucoma (n = 24), chronic angle-closure glaucoma (n = 15), and congenital/juvenile glaucoma (n = 5). The mean pre-operative intraocular pressure was 25.1 ± 5.3 mm Hg and the mean post-operative intraocular pressure at 12 months was 17.5 ± 5.1 mm Hg (p = 0.004). The mean number of intraocular pressure-lowering medications used preoperatively was 3.0 ± 1.1; the mean number of medications used at the 12-month post-operative visit was 1.4 ± 1.0 (p = 0.03). Success with one treatment was achieved in 73 (76.8%) of patients. With multiple treatments, all patients had significant intraocular pressure-lowering compared to baseline. The maximum number of treatments received by any single patient was 5. There were no instances of prolonged intraocular inflammation or long-term hypotony. CONCLUSION: Micropulse transscleral cyclophotocoagulation appears to be a safe and efficacious treatment for glaucoma. Given its improved safety profile compared to continuous-wave transscleral cyclophotocoagulation, it deserves consideration as a primary procedure.

Balanitis xerotica obliterans: an update for clinicians.

Lichen sclerosus (LS) is a severe, chronic, dermatosis characterised by inflammatory, sclerotic, pruritic lesions that causes significant morbidity in patients of all genders and ages. In boys, the lesions typically affect the foreskin and glans (termed balanitis xerotica obliterans (BXO)), leading to phimosis and potentially meatal stenosis. The incidence of the disease is not well reported but the average age of affected boys is 8 years (range 1-16). Diagnosis can often be made clinically, although histological study remains important to rule out important differential diagnoses. Complications include genital scarring, urinary and sexual dysfunction as well as the development of carcinomas in adult life. Circumcision has been regarded as definitive management of BXO in boys, but this may be supplemented with medical therapies such as topical steroids, immune modulators, intralesional triamcinolone and ozonated olive oil. Supportive measures including emollients, avoidance of irritants, surveillance of complications and recurrence as well as education and counselling remain important.Conclusion: BXO remains an important cause of phimosis in boys. The frequency of this condition appears unclear but seems likely to be less than 1% of males. Treatment generally involves circumcision, with some evidence that topical steroids or immunomodulators may decrease the incidence of recurrent meatal stenosis.What is Known:• Surgical circumcision is considered the definitive management of BXO• Many aspects of BXO are still in contention or require further study including the epidemiology and aetiology.What is New:• There is increasing awareness of non-surgical modalities that may be used in adjunct to surgery including topical corticosteroids, immune modulators, intralesional triamcinolone and ozonated olive oil• Awareness of meatal stenosis-related BXO has led to the development of surgical techniques such as preputioplasty as well as buccal mucosal inlay grafts.

Detecting Glaucoma Progression Using Guided Progression Analysis with OCT and Visual Field Assessment in Eyes Classified by International Classification of Disease Severity Codes.

PURPOSE: To compare the detection and rates of progressive retinal nerve fiber layer (RNFL) and ganglion cell-inner plexiform layer (GCIPL) loss with spectral-domain (SD) OCT and visual field (VF) loss using Guided Progression Analysis (GPA; Carl Zeiss Meditec, Dublin, CA) in glaucomatous eyes classified using International Classification of Diseases (ICD) diagnosis codes. DESIGN: Retrospective cohort study. PARTICIPANTS: Glaucoma patients with at least 3 years of follow-up and a minimum of 4 SD OCT and 5 reliable VF examinations. METHODS: Glaucoma severity was classified using ICD, 10th Edition, Clinical Modification, diagnosis codes. Rates of RNFL, macular GCIPL, and VF loss were calculated, and progression estimates were compared using generalized estimating equations and McNemar's tests. MAIN OUTCOME MEASURES: Progressive RNFL, GCIPL, and VF loss assessed by GPA criteria. RESULTS: A total of 147 eyes of 116 patients (mean age, 69.9±8.5 years) were included with mean follow-up of 69.67±18.64 months. Overall, 38 of 147 eyes (25.9%) showed RNFL progression, 35 eyes (23.8%) showed GCIPL progression, and 20 eyes (13.6%) showed VF progression. Progression by all 3 methods was noted in 10 eyes (7.0%). Eyes with mild (n = 62) and severe (n = 46) glaucoma showed significantly more progression on SD OCT compared with VF (P < 0.001 and P = 0.004). Retinal nerve fiber layer, GCIPL, and VF progressors showed faster rates of loss in average RNFL, GCIPL, and VF mean deviation compared with nonprogressors (mean ± standard error: -1.47±0.30 µm/year vs. -0.03±0.12 µm/year [P = 0.0001], -1.68±0.34 µm/year vs. -0.29±0.07 µm/year [P = 0.0001], and -1.07±0.20 dB/year vs. -0.19±0.04 dB/year [P = 0.0001], respectively). CONCLUSIONS: Spectral-domain OCT progression was significantly more common than VF progression in glaucomatous eyes classified with mild and severe disease. Structure and function should be monitored closely across the entire spectrum of glaucoma severity.

Outcomes following Autologous Fat Grafting for Oncologic Head and Neck Reconstruction.

BACKGROUND: The aims of this study were to examine the surgical, oncologic, and aesthetic outcomes of patients undergoing autologous fat grafting for oncologic head and neck reconstruction. METHODS: A review was performed of 119 consecutive patients undergoing autologous fat grafting for oncologic head and neck reconstruction from 2005 to 2014. Aesthetic outcomes were rated by a group of 10 plastic surgeons and 10 laypersons using a five-point Likert scale. RESULTS: A total of 190 fat grafting procedures were performed in 116 patients. Of these, 69 percent had received radiation therapy before their reconstructive surgery. The average number of treatments per patient was 1.6 ± 1 (range, 1 to 6), with a mean injection volume of 24.8 ± 20.2 ml. The mean follow-up time from the first fat grafting procedure was 35.8 ± 23.1 months. Complications occurred in 6 percent, none of which required a return to the operating room or hospital admission. Oncologic recurrence was observed in six patients (three local, one regional, and two distant). Locoregional recurrences were in areas outside of where the fat was grafted. Aesthetic analysis revealed that both plastic surgeons and laypersons thought patients appeared closer to normal following autologous fat grafting. CONCLUSION: This study suggests that autologous fat grafting plays a valuable role in enhancing aesthetic outcomes either by complementing or replacing reconstructive flaps for oncologic head and neck reconstruction while having both low complication rates and no evidence of being associated with cancer recurrence.

The complications associated with Resuscitative Endovascular Balloon Occlusion of the Aorta (REBOA).

Non-compressible torso hemorrhage (NCTH) remains a significant cause of morbidity and mortality in the field of trauma and emergency medicine. In recent times, there has been a resurgence in the adoption of Resuscitative Endovascular Balloon Occlusion of the Aorta (REBOA) for patients who present with NCTH. Like all medical procedures, there are benefits and risks associated with the REBOA technique. However, in the case of REBOA, these complications are not unanimously agreed upon with varying viewpoints and studies. This article aims to review the current knowledge surrounding the complications of the REBOA technique at each step of its application.

Resuscitative endovascular balloon occlusion of the aorta (REBOA): an updated review.

In a current scenario where trauma injury and its consequences account for 9% of the worlds causes of death, the management of non-compressible torso hemorrhage can be problematic. With the improvement of medicine, the approach of these patients must be accurate and immediate so that the consequences may be minimal. Therefore, aiming the ideal method, studies have led to the development of Resuscitative Endovascular Balloon Occlusion of the Aorta (REBOA). This procedure has been used at select trauma centers as a resuscitative adjunct for trauma patients with non-compressible torso hemorrhage. Although the use of this technique is increasing, its effectiveness is still not clear. This article aims, through a detailed review, to inform an updated view about this procedure, its technique, variations, benefits, limitations and future.

Resuscitative endovascular balloon occlusion of the aorta (REBOA): an updated review / Oclusão ressuscitativa por meio de balão endovascular da aorta (REBOA): revisão atualizada

ABSTRACT In a current scenario where trauma injury and its consequences account for 9% of the worlds causes of death, the management of non-compressible torso hemorrhage can be problematic. With the improvement of medicine, the approach of these patients must be accurate and immediate so that the consequences may be minimal. Therefore, aiming the ideal method, studies have led to the development of Resuscitative Endovascular Balloon Occlusion of the Aorta (REBOA). This procedure has been used at select trauma centers as a resuscitative adjunct for trauma patients with non-compressible torso hemorrhage. Although the use of this technique is increasing, its effectiveness is still not clear. This article aims, through a detailed review, to inform an updated view about this procedure, its technique, variations, benefits, limitations and future.

RESUMO Em um cenário atual onde a lesão traumática e suas consequências representam 9% das causas de morte no mundo, o manejo da hemorragia não compressível do tronco pode ser problemático. Com a melhoria da medicina, a abordagem desses pacientes deve ser precisa e imediata, para que as consequências possam ser mínimas. Portanto, visando o método ideal de manejo, estudos levaram ao desenvolvimento da técnica de oclusão ressuscitativa por balão endovascular da aorta (Resuscitative Endovascular Balloon Occlusion of the Aorta - REBOA). Este procedimento foi utilizado em centros de trauma selecionados como um complemento durante a reanimação para pacientes vítimas de trauma com hemorragia não compressível do tronco. Embora o uso dessa técnica esteja aumentando, sua eficácia ainda não é clara. Este artigo objetiva, por meio de uma revisão detalhada, trazer uma visão atualizada sobre este procedimento, sua técnica, variações, benefícios, limitações e futuro.

Genome Sequences of Chancellor, Mitti, and Wintermute, Three Subcluster K4 Phages Isolated Using Mycobacterium smegmatis mc2155.

Mycobacteriophages Chancellor, Mitti, and Wintermute infect Mycobacterium smegmatis mc2155 and are closely related to phages Cheetobro and Fionnbharth in subcluster K4. Genome sizes range from 57,697 bp to 58,046 bp. Phages are predicted to be temperate and to infect the pathogen Mycobacterium tuberculosis.

Long-term outcomes of the minimally invasive free vascularized omental lymphatic flap for the treatment of lymphedema.

BACKGROUND: The free vascularized omental lymphatic flap provides an option without the risk for iatrogenic donor site lymphedema that plagues alternative lymph node transfer donor sites. The omental flap has been associated with significant morbidity in the past; however, with modern techniques and advanced in technology, a minimally invasive approach to flap harvest is feasible. We present the long-term outcomes of the minimally invasive free vascularized omental lymphatic flap for the treatment of lymphedema. METHODS: All consecutive patients with advanced lymphedema undergoing minimally invasive free vascularized omental lymphatic flap transfer were included. Perioperative evaluation included qualitative assessments, lymphoscintigraphy, and volumetric measurements. RESULTS: Overall, 42 patients underwent a free omental lymphatic flap and had a mean follow-up of 14 (3-32) months. Subjective improvements were noted in 83% of patients. Mean volumetric improvement was 22%. Complications occurred in 16% (n = 7) of patients; this included one episode of pancreatitis and one flap loss. Postoperative imaging revealed viable lymphatic transfers. Cellulitis history was present in 74% (n = 31) patients with post-operative cellulitis occurring in 5% (n = 2) patients. CONCLUSIONS: The minimally invasive free vascularized omental lymphatic flap provides a safe donor site, a durable and versatile flap, and an efficacious therapy against lymphedema and lymphedema-related cellulitis. J. Surg. Oncol. 2017;115:84-89. © 2016 Wiley Periodicals, Inc.

Perforator Mapping and Optimizing Design of the Lateral Arm Flap: Anatomy Revisited and Clinical Experience.

BACKGROUND: The lateral arm flap remains an underused flap, especially as a free flap. In this article, the authors describe the perforator anatomy to optimize flap design and harvest. METHODS: Perforator locations were mapped in 12 cadavers (24 arms), and a retrospective review was conducted of 51 patients undergoing lateral arm flap surgery. RESULTS: One to three reliable perforators supply the lateral arm flap. Based on cadaveric dissections, from the deltoid insertion, the A, B, and C perforators were located at 7.2 ± 1.0 cm, 9.9 ± 1.2 cm, and 11.8 ± 0.8 cm, which was 0.44, 0.61, and 0.72 of the distance from the deltoid insertion, respectively. The average pedicle length was 7.0 ± 1.1 cm. The cadavers were entirely symmetric in the number and location of the perforators between the right and left arms. All 51 patients (24 male and 27 female patients) had at least one perforator with an average pedicle length of 7.0 ± 1.3 cm, an average arterial diameter of 1.7 ± 0.3 mm, and a vein diameter of 2.5 ± 0.5 mm. All but one flap was performed as a free flap for head and neck reconstruction, with one pedicled flap for shoulder reconstruction. The average flap size was 72.2 ± 37.1 cm (range, 21 to 165 cm). The nondominant arm was used for all free flaps. There were no total or partial flap losses. Twenty-eight patients reported donor-site numbness, with one infection, one hematoma, and one wound dehiscence. CONCLUSION: The lateral arm flap can be harvested reliably based on well-defined perforators and anatomical landmarks with minimal donor-site morbidity and should be included among the techniques used by reconstructive microsurgeons. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

Locus Coeruleus and Tuberomammillary Nuclei Ablations Attenuate Hypocretin/Orexin Antagonist-Mediated REM Sleep.

Hypocretin 1 and 2 (Hcrts; also known as orexin A and B), excitatory neuropeptides synthesized in cells located in the tuberal hypothalamus, play a central role in the control of arousal. Hcrt inputs to the locus coeruleus norepinephrine (LC NE) system and the posterior hypothalamic histaminergic tuberomammillary nuclei (TMN HA) are important efferent pathways for Hcrt-induced wakefulness. The LC expresses Hcrt receptor 1 (HcrtR1), whereas HcrtR2 is found in the TMN. Although the dual Hcrt/orexin receptor antagonist almorexant (ALM) decreases wakefulness and increases NREM and REM sleep time, the neural circuitry that mediates these effects is currently unknown. To test the hypothesis that ALM induces sleep by selectively disfacilitating subcortical wake-promoting populations, we ablated LC NE neurons (LCx) or TMN HA neurons (TMNx) in rats using cell-type-specific saporin conjugates and evaluated sleep/wake following treatment with ALM and the GABAA receptor modulator zolpidem (ZOL). Both LCx and TMNx attenuated the promotion of REM sleep by ALM without affecting ALM-mediated increases in NREM sleep. Thus, eliminating either HcrtR1 signaling in the LC or HcrtR2 signaling in the TMN yields similar effects on ALM-induced REM sleep without affecting NREM sleep time. In contrast, neither lesion altered ZOL efficacy on any measure of sleep-wake regulation. These results contrast with those of a previous study in which ablation of basal forebrain cholinergic neurons attenuated ALM-induced increases in NREM sleep time without affecting REM sleep, indicating that Hcrt neurotransmission influences distinct aspects of NREM and REM sleep at different locations in the sleep-wake regulatory network.