bluuebunny/crossref_metadata_2025_split huggingface dataset:
1522-P: Identification of AgRP Cells in the Murine Hindbrain That Drive Feeding
Authors: BACHOR TOMAS P. , ATTAL KUSH , VAGENA EIRINI , MIFSUD FRANCOIS , PHAM VIANA , VALDEARCOS-CONTRERAS MARTIN , VAISSE CHRISTIAN , XU ALLISON 
Published: June 2023

The central melanocortin system is essential for the regulation of food intake and body weight in humans and rodents. Agouti-related protein (AgRP) is the sole orexigenic component of the central melanocortin system and is conserved across mammalian species. AgRP is currently known to express exclusively in the mediobasal hypothalamus, and hypothalamic AgRP-expressing neurons are essential for feeding regulation. Here we describe a previously unknown population of AgRP cells in the area postrema (AP) and the adjacent commissural nucleus of the solitary tract (cNTS) of the causal brainstem. AgRP was expressed in the embryonic AP, and hindbrain AgRP expression in adult mice was low under ad libitum fed condition but increased with food deprivation. AgRP cells in the AP and cNTS consisted of locally projecting neurons as well as tanycyte-like cells. In mice that lack hypothalamic AgRP neurons, diphtheria-toxin mediated ablation of AgRP cells led to anorexia and weight loss, whereas chemogenetic activation of AgRP neurons resulted in hyperphagia and weight gain. Moreover, focal transcranial photo-stimulation of AgRP cells in the AP and cNTS with a step-function opsin with ultra-high light sensitivity (SOUL) stimulated feeding in mice with or without hypothalamic AgRP neurons, suggesting that the hyperphagic effects of hindbrain AgRP neurons are independent of hypothalamic AgRP neurons. Collectively, our study indicates that the hindbrain has a balanced melanocortin system, complete with both orexigenic and anorexigenic components, and that AgRP neurons in the hindbrain drive feeding in adult animals.

Disclosure
T.P.Bachor: None. K.Attal: None. E.Vagena: None. F.Mifsud: None. V.Pham: None. M.Valdearcos-contreras: None. C.Vaisse: None. A.Xu: Advisory Panel; Pennington Biomedical Research Center, Research Support; Eli Lilly and Company.

Funding
National Institute of Diabetes and Digestive and Kidney Diseases (1P30DK098722-01A1, P30DK63720-06A1)
1524-P: Selective Deletion of Nav1.7 in Cerebellar Purkinje Neurons Prevents Diet-Induced Obesity by Promoting Voluntary Locomotion
Authors: FENG BING , GAO PEIYU , HARMS JERNEY , LUO PEI , XU PINGWEN , HE YANLIN 
Published: June 2023

The pacemaker activity of cerebellum Purkinje neurons is well-known to be essential in multiple physiological functions, including voluntary locomotion, balance, aggressive behavior, and cognitive behavior. However, the specific ionic mechanism of Purkinje neurons in regulating voluntary locomotion is still unclear. To this end, we ex vivo electrophysiologically recorded the activity changes of Purkinje neurons after wheel running by using Pcp2-Cre/Rosa26-LTL-tdTOMATO reporter mice with tdTOMATO selectively expressed in Purkinje neurons. We found that the Purkinje neural activity was reduced by wheel running, associated with decreases in voltage-gated sodium current and expression of the voltage-gated sodium ion channel Nav1.7 (Scn9a), suggesting a potential role of Nav1.7-mediated sodium current. To test whether Nav1.7 is required for the activity dynamics of Purkinje neurons induced by voluntary wheel running and subsequent beneficial metabolic effects, we crossed Scn9a flox/flox mice with Pcp2-Cre mice to generate a mouse model with Nav1.7 selectively deleted from Purkinje neurons (Pcp2-Cre/Scn9a flox/flox, Nav1.7-KOPcp2). Nav1.7-KOPcp2 significantly decreased Purkinje neural dynamics, mimicking wheel running-induced neural activity reduction. Nav1.7-KOPcp2 mice also significantly increased voluntary locomotion compared to the control littermates. Further, Nav1.7-KOPcp2 mice showed decreased body weight gain, reduced blood glucose, and improved insulin sensitivity when fed a high-fat diet (HFD). These beneficial metabolic effects were associated with increased running activity but not food intake.
In summary, these findings provide evidence to support the vital role of the Nav1.7 ion channel expressed by Purkinje neurons in the beneficial metabolic effects of voluntary locomotion activity.

Disclosure
B. Feng: None. P. Gao: None. J. Harms: None. P. Luo: None. P. Xu: None. Y. He: None.

Funding
National Institutes of Health (R01DK123098, R01DK129548)
1526-P: Cephalic Phase Insulin Release Requires Alpha Cells
Authors: TRIMIGLIOZZI KELLY A. , WIEDEMANN SOPHIA J. , MEIER DANIEL T. , RACHID LEILA , MEREAU HELENE , BÖNI-SCHNETZLER MARIANNE , DONATH MARC Y. 
Published: June 2023

The glucose-independent rise of insulin levels immediately at the start of a meal is known as the cephalic phase of insulin release and is a neurally mediated physiological reflex. We recently found that IL-1β mediates cephalic phase insulin secretion by modulating parasympathetic neuronal transmission. It has been established that pancreatic islets are richly innervated by the parasympathetic nervous system, yet the mechanism underlying neuronally mediated insulin release at basal glucose, as seen in cephalic phase, remain to be studied.
Methods: We assessed the role of glucagon in cephalic phase insulin secretion using a transgenic mouse model in which alpha cells are selectively ablated. Further, we isolated pancreatic islets of wild-type mice, as well as alpha cell-ablated mice, and assessed insulin and glucagon secretion at low glucose after stimulation with IL-1β, a muscarinic agonist (carbachol) and both combined.
Results: We observed that mice lacking glucagon-producing alpha cells fail to exhibit a cephalic phase insulin response. In vitro, we found that muscarinic activation stimulated glucagon secretion at basal glucose, but not insulin secretion. Co-stimulating islets with the muscarinic agonist and IL-1β stimulated insulin secretion at basal glucose, and strikingly this was not the case in alpha cell-ablated islets.
Conclusion: Cephalic phase of insulin release mediated by IL-1β stimulation of parasympathetic nerves requires pancreatic alpha cells. This beta-cell response may be mediated by glucagon.

Disclosure
K.A.Trimigliozzi: None. S.J.Wiedemann: None. D.T.Meier: None. L.Rachid: None. H.Mereau: None. M.Böni-schnetzler: None. M.Y.Donath: Consultant; Boehringer Ingelheim International GmbH, Eli Lilly and Company, Novo Nordisk.
1529-P: Hypothalamic FGF1 Regulates Systemic Glucose and Energy Homeostasis
Authors: SHIN HY EMI , MYOUNG SUH JAE 
Published: June 2023

Systemic glucose and energy homeostasis is maintained by interorgan communication networks and the hypothalamus plays a key part in this process. Many research are being conducted to investigate the hypothalamus as a therapeutic target for metabolic diseases such as diabetes and obesity. Recent studies demonstrate that a single intracerebroventricular (icv) injection of FGF1 causes sustained glucose remission in diabetic rodent models (Scarlett et al.), and further study is being conducted. However, there are limited findings about how metabolic status affects endogenous brain FGF1 expression and how brain FGF1 plays in energy homeostasis. A study published in 2012 (Jonker et al.) found severe insulin resistance and a defect in adipose tissue remodeling in HFD-fed FGF1 null-mice. However, these investigations have a limitation in that it is not obvious which tissue/organ mediates insulin resistance due to FGF1 depletion. We found that CNS-deletion of FGF1 (BKO) shows impaired glucose sensing and decreased ERK/STAT3 signaling in ventromedial hypothalamus (VMH) and arcuate nucleus (ARC). This decreased leptin sensitivity leads to decreased muscle glucose uptake following glucose intolerance. We evaluated FGF1 expression in hypothalamus in several metabolic states such as fasting-feeding or other postprandial stimuli to discover which hypothalamic area mediates endogenous FGF1 activity. Interestingly, FGF1 expression in the VMH, but not other brain regions, are increased upon feeding or glucose administration and this induction of FGF1 does not require leptin or insulin. Acute FGF1 deletion in adult VMH causes glucose intolerance as well as increased body weight gain due to reduced leptin sensitivity and brown fat dysfunction, which leads to decreased energy expenditure. In keeping with this finding, germ-line ablation of FGF1 in VMH neurons results in increased adiposity. We propose that endogenous brain FGF1 regulates glucose and energy homeostasis through modulating VMH leptin sensitivity.

Disclosure
H.Shin: None. J.Suh: None.
1532-P: A Novel Light-Responsive Adipose–Hypothalamus Axis in Metabolic Regulation
Authors: TSUJI TADATAKA , LIAN HUANG TIAN , ZHANG YANG , LYNES MATTHEW , PROFETA GERSON , NARAIN NIVEN , KIEBISH MICHAEL , TSENG YU-HUA 
Published: June 2023

Light plays an essential role in life. Animals and humans possess light-sensing photoreceptors in visual and non-visual organs. We and others recently showed a non-visual role for Opsin3 (Opn3) in regulating adipose function. In this study, we discovered that direct blue light exposure to subcutaneous white adipose tissue (WAT) could reduce lipids accumulation in WAT and improve high-fat diet-induced metabolic abnormalities in an Opn3-dependent manner. Adipose tissue is an orchestrated matrix comprising high innervation that can engage in crosstalk with other organs. Particularly, it communicates with the brain via the sympathetic nervous system (SNS). Using the metabolomic approach, we found that blue light increased circulating levels of histidine, which then triggered the activation of histaminergic neurons in the hypothalamus and turned on brown adipose tissue via SNS. The administration of histidine decarboxylase antagonist blunted the effects of blue light. Taken together, these data demonstrate a novel light-responsive adipose-hypothalamus axis in metabolic regulation and provide a potential light-based therapeutic approach for obesity.

Disclosure
T.Tsuji: None. T.Huang: None. Y.Zhang: None. M.Lynes: None. G.Profeta: None. N.Narain: None. M.Kiebish: None. Y.Tseng: Consultant; Cellarity, LyGenesis.

Funding
American Heart Association (903968)
1533-P: Mind over Metabolism—A Molecular Subtype of Vagal Neurons That Can Improve Glucose Tolerance
Authors: CONLEY NICHOLAS J. , KAUFFMAN LILY S. , CAMPBELL JOHN 
Published: June 2023

Parasympathetic neurons of the dorsal motor nucleus of the vagus (DMV) control glucose metabolism through the vagus nerve and so are potential targets for diabetes treatment. Currently, little is known about the molecular identity or organization of the glucose-regulating DMV neurons. To identify these neurons for further study, we optogenetically activated them in awake mice during a glucose tolerance test (GTT). We transgenically targeted a blue light-sensitive ion channel, CaTCh, selectively to all DMV neurons based on co-expression of Chat - and Phox2b- driven recombinases (Chat-Cre::Phox2b-Flp::CaTCh mice). We implanted optical fibers over the DMV in these adult mice (2 females, 1 male), acclimated them to the test environment and procedure, and then fasted them overnight. After recording baseline glucose levels, we administered intraperitoneal glucose (20% glucose, 10uL/g) and concurrently began photostimulation (time=0-30min; 5Hz, 1ms pulse, 2.5s off, 1s on) or sham stimulation. We performed tail blood glucometry at 5-min increments from -20-120min and compared average readings between stimulation and sham trials. Results: sham 0-30min 472.6 ± 74mg/dL, stim 0-30min 352 ± 48mg/dL (p=0.0005, paired t-test); other timepoints, p&gt;0.05 between conditions. Our results indicate that activating DMV neurons can significantly improve glucose tolerance. To identify specific DMV neurons controlling glucose tolerance, we optogenetically activated a molecular subtype of DMV neurons defined by co-expression of Calb2 and Chat (2 female Calb2-Cre::Chat-Flp::CaTCh adult mice). We photostimulated Calb2+ DMV neurons while performing GTT as described above. Results: sham 0-30min 493 ± 90mg/dL, stim 0-30min 407 ± 69mg/dL (p=0.0390, paired t-test); other timepoints, p&gt;0.05 between conditions. Together our data indicates that activating DMV neurons generally or Calb2+ DMV neurons specifically improves glucose tolerance.

Disclosure
N.J.Conley: None. L.S.Kauffman: None. J.Campbell: None.

Funding
American Diabetes Association (1-18-INI-14 to J.C.)
1535-P: Physiological Role of Stress-Activated Neurons in the Paraventricular Hypothalamus
Authors: ESPINOZA DIEGO G. , ALVARSSON ALEXANDRA , JIMENEZ GONZALEZ MARIA , HAMPTON ROLLIE F. , LI ROSEMARY Y. , DEVARAKONDA KAVYA , CHOUDHURY JESLYN , WAXMAN TALIA S. , STANLEY SARAH 
Published: June 2023

Psychological stimuli that disrupt homeostasis - stressors - lead to acute and chronic metabolic disturbance but the underlying neural circuits are incompletely understood. The paraventricular nucleus of the hypothalamus (PVH) plays a significant role in behavioral, autonomic and hormonal responses to stress. The PVH contains multiple cell types including neurons expressing tyrosine hydroxylase (TH). However, the contributions of PVHTH neurons to the acute stress response are largely unknown. We aim to determine the roles of PVHTH neurons in the metabolic and behavioral responses to acute stress.
We first generated mice with tdTtomato expression in TH+ neurons (TH-tdTomato) by crossing validated TH-cre mice to mice with cre-dependent expression of tdTomato. Using immunohistochemistry for PVH neuropeptides linked to energy metabolism and stress in TH-Tdtomato mice, we quantified overlap of PVHTH neurons with CRH, oxytocin and vasopressin. We found small fractions of PVHTH neurons co-express CRH, oxytocin and vasopressin suggesting PVHTH neurons are a distinct population. Next, we assessed activation of PVHTH in response to acute stress by examining Fos expression in TH-tdTomato mice. Acute stress significantly increased the number of FOS+ PVHTH neurons. Finally, we examined whether chemogenetic activation of PVHTH neurons mimicked behavioral, hormonal and metabolic adaptations to stress using PVH injection of AAV with cre-dependent hM3DGq-mCherry or mCherry (control) in TH-cre mice. Chemogenetic activation of PVHTH neurons acutely suppressed dark-phase food intake and impaired glucose tolerance without effects on plasma corticosterone or anxiety-like behavior.
These findings suggest acute stress may recruit PVHTH neurons to elicit hypophagia and impair glucose tolerance, independent of HPA axis activation and anxiety-like behavior. Future studies using chemogenetic inhibition will determine if PVHTH neurons are required for the full metabolic response to acute stress.

Disclosure
D.G.Espinoza: None. A.Alvarsson: None. M.Jimenez gonzalez: None. R.F.Hampton: None. R.Y.Li: None. K.Devarakonda: None. J.Choudhury: None. T.S.Waxman: None. S.Stanley: Consultant; Redpin Therapeutics.
1536-P: Effects of Sucralose vs. Sucrose on Hypothalamic Blood Flow, Glucose Signaling, and Appetite
Authors: CHAKRAVARTTI SANDHYA P. , KULLMANN STEPHANIE , VEIT RALF , YUNKER ALEXANDRA G. , ANGELO BRENDAN C. , JANN KAY , LUO SHAN H. , XIANG ANNY , MONTEROSSO JOHN R. , PAGE KATHLEEN A. 
Published: June 2023

Sucralose is a non-nutritive sweetener (NNS) commonly consumed to provide sweet taste without calories. Yet, some studies suggest NNS stimulate appetite, potentially by altering post-ingestive metabolic signals that communicate with the hypothalamus to reduce hunger. We tested the hypothesis that sucralose vs sucrose would elicit different metabolic, hypothalamic blood flow (BF) and appetite responses, and that post-ingestive increases in metabolic signals would be associated with reductions in hypothalamic BF and hunger. In a randomized cross-over trial, hypothalamic BF (measured by perfusion MRI), appetite ratings, and glucose/hormone responses were obtained before, 15 and 30 min after ingestion of drinks containing sucrose (75g) or sucralose (sweetness matched). Paired t-tests were used to compare sucralose vs sucrose on metabolic responses, hypothalamic BF, and appetite. Linear regression was used to examine associations between peripheral glucose, insulin, GLP-1 and hypothalamic BF and hunger. 76 adults (43F; 23.3±3.9 yrs; BMI 27.2±5) completed the study. Subjects reported increased hunger at 15 (p&lt;.04) and 30min (p&lt;.003) after sucralose vs sucrose. Peripheral glucose, insulin, and GLP-1 were lower after sucralose vs sucrose (all p&lt;.0001). Glycemic response (whole hypothalamus (β=-.002, p=.05), medial subfield (β=−.005, p&lt;.006)), but not insulin or GLP-1, was associated with the hypothalamic response to sucrose at 30 min, controlling for age, sex, BMI. These associations were not observed after sucralose (β=.008 p=.2). The medial hypothalamic response predicted hunger response after both drinks (β=1.79, p&lt;.02). Sucralose vs sucrose elicited disparate metabolic, hypothalamic, and hunger responses. Sucrose-induced increases in glycemia were associated with reductions in medial hypothalamic BF, which was linked to reduced hunger. Compared to sucrose, post-ingestive signaling responses were altered after sucralose, which may affect appetite.

Disclosure
S.P.Chakravartti: None. K.A.Page: None. S.Kullmann: None. R.Veit: None. A.G.Yunker: None. B.C.Angelo: None. K.Jann: None. S.H.Luo: None. A.Xiang: None. J.R.Monterosso: None.

Funding
National Institute of Diabetes and Digestive and Kidney Diseases (R01DK102794)
1538-P: Brain TGFß Regulates Glucose Tolerance in Male Mice with Metabolic Syndrome
Authors: ELGAZZAZ MONA , MIAO KEVIN , NUZZO ANNA R. , RESTIVO LUKE , MOLES KARL , LAZARTIGUES ERIC D. 
Published: June 2023

Hypothalamic inflammation contributes to insulin resistance and obesity. Transforming growth factor beta (TGFβ) is among the inflammatory mediators inducing hypothalamic inflammation. In this study, we investigated the role of brain TGFβ on metabolic function. We hypothesized that TGFβ in the brain promotes insulin resistance and reduces glucose tolerance in diabetic and obese mice. C57BL/J male mice were fed either hypercaloric diet (HCD, 52% fat) or regular diet (RD) from weaning. At 3 months age, intraperitoneal glucose tolerance test (GTT), insulin tolerance test (ITT) and weight confirmed the acquisition of metabolic syndrome. Hypothalamic protein expression was assessed for TGFβ receptor 2, phosphorylated SMAD3, SMAD6 and SMAD7, as downstream signal transducer proteins for TGFβ. Compared to RD male mice (n=6), mice on HCD (n=6) showed glucose intolerance (AUC; 60587 ±2460 vs. 30802±1923, p&lt;0.0001), insulin intolerance (AUC; Mean±SEM; 19025836.0 vs.11436±1087, p=0.02) and weight gain (33.67±2.028 vs. 26.89±0.4231 g). HCD mice also exhibited increased protein levels of TGFβR2 (normalized to total protein, Fold Change (FC)±SEM to RD group; 3.54±0.3, p&lt;0.0001), pSMAD3 (FC 2±0.5, p=0.04) while inhibitors for TGFβ signaling such as SMAD6 and SMAD7, were downregulated (FC 0.7±0.1, p=0.03 and 0.8±0.04, p=0.03, respectively). To understand the role of brain TGFβ on metabolic regulation, 3 months HCD mice were subjected to intracerebroventricular (ICV) infusion with either TGFβ neutralizing antibody (nAb, n=4) or IgG control antibody (n=4) using subcutaneous osmotic pumps connected to ICV cannula for 2 weeks. GTT was measured after 2-week infusion. Treatment with TGFβ nAb resulted in lower fasting blood glucose compared to before treatment (166.1 ±6.878 vs. 229.9 ±3.923 mg/dL) and at 120 min time point (229.9 ±28.69 vs. 382.2 ±24.33 mg/dL). In conclusion, brain TGFβ plays a key role in regulating glucose metabolism probably through inflammation-induced insulin resistance.

Disclosure
M.Elgazzaz: None. K.Miao: None. A.R.Nuzzo: None. L.Restivo: None. K.Moles: None. E.D.Lazartigues: None.

Funding
American Diabetes Association (1-19-IBS-291 to E.D.L.)
1539-P: Effects of Free Fatty Acids (FFA) on Central Regulation of Glucose Metabolism
Authors: FABIOLA NOGUEIRA CORDEIRO MONICA , ROUTH VANESSA H. , MANAVALAN ANJALI , MATHIAS PRIYANKA M. , BOYLE CAROLINE E. , RIOS CHEN ANA C. , JIANG NANCY , FRIEDMAN NAOMI , EMRE DEMIRHAN YUNUS , HAWKINS MEREDITH , ZHANG KEHAO 
Published: June 2023

Activation of hypothalamic KATP channels reduces endogenous glucose production (EGP) in nondiabetic rodents and humans, but this response is lost in T2D. Lowering FFA with nicotinic acid (NA) restored central regulation of EGP in T2D subjects (Diabetes 2019; 1799P). High FFA levels, typical of T2D, may increase endoplasmic reticulum (ER) stress and thereby impair hypothalamic control of EGP.
To explore how increased FFA might impact central regulation of EGP, we harvested hypothalamic wedges from n=10 Zucker Diabetic Fatty (ZDF) rats (glucose ~ 370 mg/dL) following infusion of NA or saline (Alzet minipump) for 18 h. Nondiabetic rats (n=5) were studied as controls.
Gene expression of KATP channel subunit Kir6.2 was decreased in ZDF rats. Upon lowering FFA with NA, hypothalamic expression of this major subunit was restored (Fig 1a). Conversely, increased expression of genes regulating ER stress was reversed by NA (Fig 1b). Directly treating ZDF rat brain slices with NA did not alter the expression of these genes, suggesting that NA's effects were mediated by lowering FFA (Fig 1c).
In conclusion, hypothalamic effects of elevated FFA include reduced KATPchannel expression and increased expression of ER stress genes, both of which may contribute reversibly to the loss of central regulation of EGP in T2D. These findings are consistent with our observations in humans noted above, suggesting that FFA could be a therapeutic target in T2D.

Disclosure
M. Nogueira Cordeiro: None. V.H. Routh: None. A. Manavalan: None. P.M. Mathias: None. C.E. Boyle: None. A.C. Rios Chen: None. N. Jiang: None. N. Friedman: None. Y. Demirhan: None. M. Hawkins: None. K. Zhang: None.

Funding
American Diabetes Association (7-08-CR-28 to M.H.); National Institutes of Health (R01DK069861); Diabetes Action Research and Education Foundation
1540-P: Diazoxide (DZ) May Lower Endogenous Glucose Production (EGP) via Mitochondrial ATP-Sensitive K+ (mt KATP) Channels
Authors: VAIL GWYNDOLIN , PATEL VISHWENDRA , SARKAR PALLABI , ZHANG KEHAO , HAWKINS MEREDITH , ROUTH VANESSA H. 
Published: June 2023

Rodent and human studies suggest that ventromedial hypothalamus (VMH) KATP channels suppress EGP. The VMH possesses glucose-sensing neurons that are either excited (GE) or inhibited (GI) by glucose. Since administration of the KATP channel activator DZ reduces EGP in rodents and humans, we examined the impact of DZ on VMH glucose sensing neurons. DZ opens both plasma membrane and mt KATP channels; the former inhibits neurons while the latter maintains mitochondrial metabolic function. We hypothesize that the concentration of brain DZ that lowers EGP affects the mt and not the plasma membrane KATP channel of VMH glucose sensing neurons. VMH glucose-excited (GE) neurons were exposed to 0.1 µM DZ in 2.5 mM glucose for 20 minutes prior to lowering glucose to 0.1 mM in the presence of DZ. DZ did not alter membrane potential (MP; 2.5 glucose: −63 ± 4 mV; DZ: −64 ± 5 mV, P = 0.3807, N = 7) or cell resistance (R; 2.5 glucose: 658 ± 136 MΩ; DZ 656 ± 146 MΩ, P = 0.9526, N = 7) in 2.5 mM glucose. There was also no difference in the MP or R response to lowering glucose from 2.5 to 0.1 mM in the presence or absence of DZ (% change MP control: −20.8 ± 3, diazoxide: −19.9 ± 3, P = 0.9044, N = 5; R control: −59.0 ± 5, diazoxide: −55.3 ± 6, P = 0.7134, N = 5). Similar results were seen with VMH glucose-inhibited neurons (N=4; Fig. 1). Our results are consistent with an action of DZ on mt but not plasma membrane KATP channels of VMH glucose sensing neurons.

Disclosure
G.Vail: None. V.Patel: None. P.Sarkar: None. K.Zhang: None. M.Hawkins: None. V.H.Routh: None.

Funding
National Institutes of Health (2R01DK069681-11)