Neuromodulation of orexin neurons reduces diet-induced adiposity.

BACKGROUND/OBJECTIVES: Low levels of orexin are associated with obesity and reduced physical activity in humans and animals. SUBJECTS/METHODS: Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) selectively activated orexin neurons in mouse lateral hypothalamus (LH) to measure effects on spontaneous physical activity (SPA). DREADD targeting was achieved by stereotaxic injection of AAV vectors into caudal lateral LH of heterozygous orexin-Cre or C57/B6J mice. In one set of studies, excitation of orexin neurons was examined (virus: AAV2-EF1a-DIO-hM3Dq-mCherry), and test sessions began 3-4 h after light cycle onset. In a study examining the inhibition of orexin neurons (virus: AAV2-hSyn-DIO-hM4Di-mCherry), testing began 15 min prior to dark cycle onset. Clozapine n-oxide (CNO; 1 or 5 mg/kg) or saline was injected intraperitoneally and time spent moving in open field chambers was recorded for 2 h. Follow-up studies in separate mouse cohorts quantified SPA in parallel with changes in energy expenditure (EE) and chow intake using indirect calorimetry chambers (SableSystem™). Following acclimation, testing sessions (saline and/or CNO) took place over the course of ~1 week, with injections administered every day. Changes in SPA, EE, chow intake, fecal boli, and body composition (EchoMRI™) were measured. Additional mice cohorts were fed a high-fat diet (HFD) and injected with CNO daily up to 10 days to assess the potential for orexin activation to prevent diet-induced obesity. RESULTS: Activation of orexin resulted in increases in SPA in male and female mice, and was accompanied by increases in energy expenditure without changes in overall chow intake. When orexin activation occurred in the context of high fat diet, weight gain and adiposity were significantly attenuated. SPA was decreased when DREADDs were used to inhibit orexin activity. CONCLUSION: These results demonstrate that orexin neurons play a critical role in mediating physical activity and suggest a novel therapeutic target for treating obesity.

Orexin activation counteracts decreases in nonexercise activity thermogenesis (NEAT) caused by high-fat diet.

Overweight and obesity result from an imbalance between caloric intake and energy expenditure, including expenditure from spontaneous physical activity (SPA). Changes in SPA and resulting changes in non-exercise activity thermogenesis (NEAT) likely interact with diet to influence risk for obesity. However, previous research on the relationship between diet, physical activity, and energy expenditure has been mixed. The neuropeptide orexin is a driver of SPA, and orexin neuron activity can be manipulated using DREADDs (Designer Receptors Exclusively Activated by Designer Drugs). We hypothesized that HFD decreases SPA and NEAT, and that DREADD-mediated activation of orexin neuron signaling would abolish this decrease and produce an increase in NEAT instead. To test these ideas, we characterized behaviors to determine the extent to which access to a high-fat diet (HFD) influences the proportion and probability of engaging in food intake and activity. We then measured NEAT following access to HFD and following a DREADD intervention targeting orexin neurons. Two cohorts of orexin-cre male mice were injected with an excitatory DREADD virus into the caudal hypothalamus, where orexin neurons are concentrated. Mice were then housed in continuous metabolic phenotyping cages (Sable Promethion). Food intake, indirect calorimetry, and SPA were automatically measured every second. For cohort 1 (n=8), animals were given access to chow, then switched to HFD. For cohort 2 (n=4/group), half of the animals were given access to HFD, the other access to chow. Then, among animals on HFD, orexin neurons were activated following injections of clozapine n-oxide (CNO). Mice on HFD spent significantly less time eating (p<0.01) and more time inactive compared to mice on chow (p<0.01). Following a meal, mice on HFD were significantly more likely to engage in periods of inactivity compared to those on chow (p<0.05). NEAT was decreased in animals on HFD, and was increased to the NEAT level of control animals following activation of orexin neurons with DREADDs. Food intake (kilocalories) was not significantly different between mice on chow and HFD, yet mice on chow expended more energy per unit of SPA, relative to that in mice consuming HFD. These results suggest that HFD consumption reduces SPA and NEAT, and increases inactivity following a meal. Together, the data suggest a change in the efficiency of energy expenditure based upon diet, such that SPA during HFD burns fewer calories compared to SPA on a standard chow diet.

Role of orexin-A in the ventrolateral preoptic area on components of total energy expenditure.

BACKGROUND: Identifying whether components of total energy expenditure (EE) are affected by orexin receptor (OXR1 and OXR2) stimulation or antagonism with dual orexin receptor antagonists (DORAs) has relevance for obesity treatment. Orexin receptor stimulation reduces weight gain by increasing total EE and EE during spontaneous physical activity (SPA). OBJECTIVE: The purpose of this study was to determine if a DORA (TCS-1102) in the ventrolateral preoptic area (VLPO) reduced orexin-A-induced arousal, SPA, total EE and EE during sleep, rest, wake and SPA and whether the DORA alone reduced total EE and its components. We hypothesized that: (1) a DORA would reduce orexin-A induced increases in arousal, SPA, components of total EE, reductions in sleep and the EE during sleep and (2) the DORA alone would reduce baseline (non-stimulated) SPA and total EE. SUBJECTS/METHODS: Sleep, wakefulness, SPA and EE were determined after microinjection of the DORA (TCS-1102) and orexin-A in the VLPO of male Sprague-Dawley rats with a unilateral cannula targeted towards the VLPO. Individual components of total EE were determined based on time-stamped data. RESULTS: The DORA reduced orexin-A-induced increases in arousal, SPA, total EE and EE during SPA, wake, rest and sleep 1 h post injection (P<0.05). Orexin-A significantly reduced sleep and significantly increased EE during sleep 1 h post injection (P<0.05). Furthermore, the DORA alone significantly reduced total EE, EE during sleep (NREM and REM) and resting EE 2 h post injection (P<0.05). CONCLUSIONS: These data suggest that orexin-A reduces weight gain by stimulating total EE through increases in EE during SPA, rest and sleep. Residual effects of the DORA alone include decreases in total EE and EE during sleep and rest, which may promote weight gain.

Mechanisms underlying obesity resistance associated with high spontaneous physical activity.

Obesity resistance due to elevated orexin signaling is accompanied by high levels of spontaneous physical activity (SPA). The behavioral and neural mechanisms underlying this observation have not been fully worked out. We determined the contribution of hypothalamic orexin receptors (OXRs) to SPA stimulated by orexin A (OXA), whether OXA-stimulated SPA was secondary to arousal and whether voluntary wheel running led to compensations in 24-h SPA. We further tested whether orexin action on dopamine one receptors (DA1R) in the substantia nigra (SN) plays an important role in the generation of SPA. To test this, SPA response was determined in lean and obese rats with cannulae targeted toward the rostral lateral hypothalamus (rLH) or SN. Sleep/wake states were also measured in rats with rLH cannula and electroencephalogram/electromyogram radiotelemetry transmitters. SPA in lean rats was more sensitive to antagonism of the OX1R and in the early response to the orexin 2 agonist. OXA increased arousal equally in lean and obese rodents, which is discordant from the greater SPA response in lean rats. Obesity-resistant rats ran more and wheel running was directly related to 24-h SPA levels. The OX1R antagonist, SB-334867-A, and the DA1R antagonist, SCH3390, in SN more effectively reduced SPA stimulated by OXA in obesity-resistant rats. These data suggest OXA-stimulated SPA is not secondary to enhanced arousal, propensity for SPA parallels inclination to run and that orexin action on dopaminergic neurons in SN may participate in the mediation of SPA and running wheel activity.

A scripted control system for autonomous hardware-timed experiments.

We present the labscript suite, an open-source experiment control system for automating shot-based experiments and their analysis. Experiments are composed as Python code, which is used to produce low-level hardware instructions. They are queued up and executed on the hardware in real time, synchronized by a pseudoclock. Experiment parameters are manipulated graphically, and analysis routines are run as new data are acquired. With this system, we can easily automate exploration of parameter spaces, including closed-loop optimization.

Vagal blocking improves glycemic control and elevated blood pressure in obese subjects with type 2 diabetes mellitus.

BACKGROUND: An active device that downregulates abdominal vagal signalling has resulted in significant weight loss in feasibility studies. OBJECTIVE: To prospectively evaluate the effect of intermittent vagal blocking (VBLOC) on weight loss, glycemic control, and blood pressure (BP) in obese subjects with DM2. METHODS: Twenty-eight subjects were implanted with a VBLOC device (Maestro Rechargeable System) at 5 centers in an open-label study. Effects on weight loss, HbA1c, fasting blood glucose, and BP were evaluated at 1 week to 12 months. RESULTS: 26 subjects (17 females/9 males, 51 ± 2 years, BMI 37 ± 1 kg/m(2), mean ± SEM) completed 12 months followup. One serious adverse event (pain at implant site) was easily resolved. At 1 week and 12 months, mean excess weight loss percentages (% EWL) were 9 ± 1% and 25 ± 4% (P < 0.0001), and HbA1c declined by 0.3 ± 0.1% and 1.0 ± 0.2% (P = 0.02, baseline 7.8 ± 0.2%). In DM2 subjects with elevated BP (n = 15), mean arterial pressure reduced by 7 ± 3 mmHg and 8 ± 3 mmHg (P = 0.04, baseline 100 ± 2 mmHg) at 1 week and 12 months. All subjects MAP decreased by 3 ± 2 mmHg (baseline 95 ± 2 mmHg) at 12 months. CONCLUSIONS: VBLOC was safe in obese DM2 subjects and associated with meaningful weight loss, early and sustained improvements in HbA1c, and reductions in BP in hypertensive DM2 subjects. This trial is registered with ClinicalTrials.gov NCT00555958.

A versatile high resolution objective for imaging quantum gases.

We present a high resolution objective lens made entirely from catalog singlets that has a numerical aperture of 0.36. It corrects for aberrations introduced by a glass window and has a long working distance of 35 mm, making it suitable for imaging objects within a vacuum system. This offers simple high resolution imaging for many in the quantum gas community. The objective achieves a resolution of 1.3 µm at the design wavelength of 780 nm, and a diffraction-limited field of view of 360 µm when imaging through a 5 mm thick window. Images of a resolution target and a pinhole show quantitative agreement with the simulated lens performance. The objective is suitable for diffraction-limited monochromatic imaging on the D2 line of all the alkalis by changing only the aperture diameter, retaining numerical apertures above 0.32. The design corrects for window thicknesses of up to 15 mm if the singlet spacings are modified.

Role of orexin receptors in obesity: from cellular to behavioral evidence.

The orexin peptides and their two receptors are involved in multiple physiological processes, including energy homeostasis, arousal, stress and reward. Higher signaling of the orexin peptides at the orexin receptors (OXR) protects against obesity, but it is less clear how their activation in different brain regions contributes to this behavioral output. This review summarizes the evidence available for a role of central OXR in energy homeostasis and their contribution to obesity. A detailed analysis of anatomical, cellular and behavioral evidence shows that modulation of energy homeostasis by the OXR is largely dependent upon anatomical and cellular context. It also shows that obesity resistance provided by activation of the OXR is distributed across multiple brain sites with site-specific actions. We suggest that understanding the role of the OXR in the development of obesity requires considering both specific mechanisms within brain regions and interactions of orexinergic input between multiple sites.

Effects of butorphanol on feeding and neuropeptide Y in the rat.

Butorphanol ([BT] an opioid receptor agonist/antagonist) is different from other opioid agonists in that a single dose of BT can elicit up to 12 g of chow intake in a satiated rat whereas most opioid agonists induce a mild feeding response (2-3 g). Here, we first examined whether the effectiveness of BT to elicit feeding was affected by dose, method of infusion and possible tachyphylaxis following administration. Secondly, we examined whether BT administration influenced hypothalamic NPY gene expression and peptide levels. A single dose administration of BT (4 mg/kg) significantly increased food intake at 2, 3 and 6 h after administration. However following repeated injections of BT at 4 mg/kg, the cumulative long-term intake of BT-treated rats did not differ from that of controls, indicating that the animals compensate for the increased feeding following BT injection by decreased feeding at a later time. An ascending dose schedule of repeated BT injections resulted in additional feeding. NPY gene expression in the ARC was influenced by how much food had been consumed, but not by BT. The amount of food consumed and the level of NPY mRNA were inversely correlated. This is consistent with NPY's role in normal feeding. BT treatment did not affect either NPY or leptin RIA levels. We conclude that the feeding produced by BT is sensitive to dose and dosing paradigm. Further, its mechanism of action does not appear to be mediated by NPY or leptin pathways.

Spontaneous physical activity protects against fat mass gain.

OBJECTIVE: It is unclear whether elevated spontaneous physical activity (SPA, very low-intensity physical activity) positively influences body composition long term. We determined whether SPA and caloric intake were differentially related to the growth curve trajectories of body weight, fat mass (FM) and fat-free mass (FFM) between obesity resistant and Sprague-Dawley rats at specific age intervals. DESIGN AND SUBJECTS: Body composition, SPA and caloric intake were measured in selectively-bred obesity-resistant and out-bred Sprague-Dawley rats from 1 to 18 months. Data from development throughout maturation were analyzed by longitudinal growth curve modeling to determine the rate and acceleration of body weight, FM- and FFM-gain. RESULTS: Obesity-resistant rats had a lower rate of FM gain overall, a lower acceleration in body weight early in life, significantly greater SPA and lower cumulative caloric intake. Greater SPA in obesity-resistant rats was significantly associated with a lower rate of FM gain overall and lower acceleration in body weight early in life. Obesity resistant rats lost less FFM compared with Sprague-Dawley rats despite that obesity-resistant rats had a lower acceleration in FFM gain early in life. Obesity-resistant rats gained less FM and more FFM per gram body weight and were less energy efficient than Sprague-Dawley rats. Caloric intake was significantly and positively related to body weight, FM and FFM gain in both groups. Circadian patterns of caloric intake were group and age-dependent. Our data demonstrate that elevated and sustained SPA during development and over the lifespan are related to the reduced the rate of FM gain and may preserve FFM. CONCLUSION: These data support the idea that SPA level is a reproducible marker that reliably predicts propensity for obesity in rats, and that elevated levels of SPA maintained during the lifespan promote a lean phenotype.

Elevated sleep quality and orexin receptor mRNA in obesity-resistant rats.

OBJECTIVE: To determine if resistance to weight gain is associated with alterations in sleep-wake states and orexin receptor gene expression. DESIGN: Three-month-old obesity-susceptible Sprague-Dawley (SD) and obesity-resistant (OR) rats were fed standard rodent chow. Sleep-wake cycle was measured by radiotelemetry and orexin receptor profiles in sleep-wake regulatory areas of the brain were quantified by quantitative reverse transcriptase-PCR. SUBJECTS: Adult male obesity-susceptible SD and selectively bred OR rats. MEASUREMENTS: Body weight, food intake, energy efficiency, percent time spent in active wake (AW), quiet wake (QW), slow-wave sleep (SWS), rapid eye movement (REM) sleep, number and mean duration of sleep-wake episodes, number of stage transitions, SWS sleep delta power and orexin receptor mRNA levels were measured. RESULTS: OR rats weighed significantly less and had lower energy efficiency than SD rats. Food intake was not different between SD and OR rats. Time spent in QW was similar between groups, and therefore AW and QW were combined and are referred to as 'wakefulness'. OR rats spent significantly more time in wakefulness and less time in SWS compared with SD rats during the 24-h recording period. Relative to SD rats, OR rats had significantly fewer sleep-wake episodes and the duration of the episodes were prolonged, indicating less fragmented sleep. Furthermore, OR rats had fewer transitions between sleep stages, which indicates that OR rats were behaviorally more stable and had more consolidated sleep than obesity-susceptible SD rats. OR rats showed lower delta power during SWS, indicating a lower sleep drive. Our results showed greater orexin receptor gene expression in sleep regulatory brain areas in OR rats. CONCLUSION: These results show that prolonged wakefulness, better sleep quality, lower sleep drive and greater orexin signaling may confer protection against obesity.

Hypocretin/orexin and energy expenditure.

The hypocretins or orexins are endogenous neuropeptides synthesized in discrete lateral, perifornical and dorsal hypothalamic neurones. These multi-functional neuropeptides modulate energy homeostasis, arousal, stress, reward, reproduction and cardiovascular function. This review summarizes the role of hypocretins in modulating non-sleep-related energy expenditure with specific focus on the augmentation of whole body energy expenditure as well as hypocretin-induced physical activity and sympathetic outflow. We compare the efficacy of hypocretin-1 and 2 on energy expenditure and evaluate whether the literature implicates hypocretin signalling though the hypocretin-1 and -2 receptor as having shared and or functionally specific physiological effects. Thus far data suggest that hypocretin-1 has a more robust stimulatory effect relative to hypocretin-2. Furthermore, hypocretin-1 receptor predominantly mediates behaviours known to influence energy expenditure. Further studies on the hypocretin-2 receptor are needed.

Intra-abdominal vagal blocking (VBLOC therapy): clinical results with a new implantable medical device.

BACKGROUND: A new medical device uses high-frequency electrical algorithms to create intermittent vagal blocking (VBLOC therapy). The aim is to assess the effects of vagal blocking on excess weight loss (EWL), safety, dietary intake, and vagal function. METHODS: An open-label, 3-center study was conducted in obese subjects (body mass index [BMI] 35-50 kg/m(2)). Electrodes were implanted laparoscopically on both vagi near the esophagogastric junction to provide electrical block. Patients were followed for 6 months for body weight, safety, electrocardiogram, dietary intake, satiation, satiety, and plasma pancreatic polypeptide (PP) response to sham feeding. To specifically assess device effects alone, no diet or exercise programs were instituted. RESULTS: Thirty-one patients (mean BMI, 41.2 +/- 1.4 kg/m(2)) received the device. Mean EWL at 4 and 12 weeks and 6 months after implant was 7.5%, 11.6%, and 14.2%, respectively (all P < .001); 25% of patients lost >25% EWL at 6 months (maximum, 36.8%). There were no deaths or device-related serious adverse events (AEs). Calorie intake decreased by >30% at 4 and 12 weeks and 6 months (all P 25 pg/mL (P = .02). Three patients had serious AEs that required brief hospitalization, 1 each for lower respiratory tract, subcutaneous implant site seroma, and Clostridium difficile diarrhea. CONCLUSIONS: Intermittent, intra-abdominal vagal blocking is associated with significant EWL and a desirable safety profile.

Neuropeptidergic mediators of spontaneous physical activity and non-exercise activity thermogenesis.

Lean individuals have high levels of spontaneous physical activity (SPA) and the energy expenditure derived from that activity, termed non-exercise activity thermogenesis or NEAT, appears to protect them from obesity. Conversely, obesity in different human populations is characterized by low levels of SPA and NEAT. Like in humans, elevated SPA in rats appears to protect against obesity: obesity-resistant rats have significantly greater SPA and NEAT than obesity-prone rats. We review the literature on brain mechanisms important in mediating SPA and NEAT. The focus is on neuropeptides, including cholecystokinin, corticotropin-releasing hormone (also known as corticotropin-releasing factor), neuromedin U, neuropeptide Y, leptin, agouti-related protein, orexin-A (also known as hypocretin-1), and ghrelin. We also review information regarding interactions between these neuropeptides and dopamine, a neurotransmitter important in mediating motor function. Finally, we present evidence that elevated signaling of pathways mediating SPA and NEAT may protect against weight gain and obesity.

Articular cartilage and changes in arthritis: matrix degradation.

While many proteases in articular cartilage have been described, current studies indicate that members of two families of metalloproteases - MMPs and the ADAMTSs - are responsible for the degradation of the major components of this tissue. Collagenases (MMPs) make the first cleavage in triple-helical collagen, allowing its further degradation by other proteases. Aggrecanases (ADAMTSs), in conjunction with other MMPs, degrade aggrecan, a component of the proteoglycan aggregate. Antineoepitope antibodies that recognize the cleavage products of collagen and aggrecan generated by these enzymes are now available and are being used to detect the sites of action and to quantitate degradation products.

Naloxone's effect on meal microstructure of sucrose and cornstarch diets.

The opioid receptor antagonist naloxone decreases consumption of high-sucrose diets but does not reduce cornstarch diet intake in energy-restricted rats. Sucrose-fed rats eat at a much higher rate, consuming more food than cornstarch-fed rats. We examined meal microstructure using an automated weighing system in food-restricted rats eating either a high-sucrose or high-cornstarch diet. Sucrose-fed rats exhibited a higher rate of eating during their first meal compared with cornstarch-fed rats (0.34 vs. 0.20 g/min, respectively). However, naloxone did not reduce eating rate in either group. Naloxone decreased the size of the first meal in both diet groups by shortening the length of the meal. Naloxone's anorectic effect was more potent in the sucrose-fed rats. These results indicate that naloxone's heightened anorectic effect on sucrose diet consumption is not "rate dependent." Naloxone's anorectic actions may be modulated by two conditions, the sensory properties of food and the energy state of the animal. Thus the elevated anorectic potency of naloxone in energy-restricted sucrose-fed rats may reflect actions on neural systems that mediate orosensory and/or postingestive signals.

The kappa-opioid antagonist GNTI reduces U50,488-, DAMGO-, and deprivation-induced feeding, but not butorphanol- and neuropeptide Y-induced feeding in rats.

Antagonists selective for either kappa- [e.g. nor-binaltorphimine (nor-BNI)] and mu- (e.g. beta-funaltrexamine) opioid receptors have previously been shown to reduce both kappa- and mu-opioid-induced feeding. In the present studies, the anorectic effects of GNTI, a newly synthesized antagonist selective for kappa-opioid receptors, were studied in rats. GNTI (0.032-0.32 nmol; i.c.v.), administered 15 min prior to food access, reduced feeding induced by the kappa-opioid agonist U50,488 (producing a 70% maximal decrease), the mu-opioid agonist DAMGO (90% maximal decrease), and 24 h acute food deprivation (60% maximal decrease). GNTI did not reduce the orexigenic effects of butorphanol, an agonist that binds to both kappa- and mu-opioid receptors, and neuropeptide Y (NPY). Taken together, these results suggest that GNTI is a potent anorectic agent and opioid antagonist in rats. Like nor-BNI, GNTI reduced feeding induced by both kappa- and mu-opioid agonists. However, unlike nor-BNI, GNTI did not alter the orexigenic effects of butorphanol or NPY. Given the selectivity of GNTI and its effectiveness in several of the present experiments, its potency, and its short duration of action compared to nor-BNI, GNTI may serve to be a useful tool to study behavioral effects mediated by kappa-opioid receptors.

Role of alpha-MSH in the regulation of consummatory behavior: immunohistochemical evidence.

Central injection of alpha-melanocyte-stimulating hormone (alpha-MSH) decreases food intake, suggesting a role for this peptide in the mediation of satiety. Inasmuch as alpha-MSH also supports the development of taste aversions under certain conditions, the nature of its influence on ingestive behavior, i.e., whether it is related to satiety or aversion, remains unclear. In the present studies, we used immunostaining, including that for c-Fos as a marker of neuronal activation, to further substantiate the physiological role for alpha-MSH in the regulation of consummatory behavior. We found that an increase in activation of alpha-MSH neurons in the arcuate nucleus coincided with meal termination. Administration of powerful aversive agents, LiCl and CuSO(4), did not stimulate alpha-MSH cells but did induce pronounced activation of oxytocin (OT) and vasopressin (VP) neurons, the final components of circuitry mediating aversion. We observed fewer Fos-positive OT/VP neurons after alpha-MSH injection into the lateral ventricle or into the hypothalamic paraventricular nucleus, treatments that cause mild or no aversion, respectively. The degree of activation of OT/VP neurons paralleled the magnitude of aversive response to a given treatment. Our data support the hypothesis that, in the arcuate nucleus, alpha-MSH acts as a satiety mediator independent from aversion-related mechanisms.

Identification of central sites involved in butorphanol-induced feeding in rats.

Butorphanol (BT), a mixed kappa- and mu-opioid receptor agonist, induces vigorous food intake in rats. Peripheral injection of BT seems to increase food intake more effectively than intracerebroventricular administration. To further elucidate the nature of BT's influence on consummatory behavior, we examined which feeding-related brain areas exhibit increased c-Fos immunoreactivity (IR) following subcutaneous injection of 4 mg/kg body weight BT, a dose known to induce a maximal orexigenic response. We also evaluated whether direct administration of BT into the forebrain regions activated by peripheral BT injection affects food intake. Peripheral BT administration induced c-Fos-IR in the hypothalamic paraventricular nucleus (PVN), central nucleus of the amygdala (CeA), and nucleus of the solitary tract (NTS). However, 0.1-30 microg BT infused into the CeA, failed to increase food intake 1, 2, and 4 h after injection. Only the highest dose of BT (30 microg) injected into the PVN increased feeding. These results suggest that the PVN, CeA, and NTS mediate the effects of peripherally-injected BT. The PVN or CeA are probably not the main target sites of immediate BT action.