Bromazepam increases the error of the time interval judgments and modulates the EEG alpha asymmetry during time estimation.

AIM: This study investigated the bromazepam effects in male subjects during the time estimation performance and EEG alpha asymmetry in electrodes associated with the frontal and motor cortex. MATERIAL AND METHODS: This is a double-blind, crossover study with a sample of 32 healthy adults under control (placebo) vs. experimental (bromazepam) during visual time-estimation task in combination with electroencephalographic analysis. RESULTS: The results demonstrated that the bromazepam increased the relative error in the 4 s, 7 s, and 9 s intervals (p = 0.001). In addition, oral bromazepam modulated the EEG alpha asymmetry in cortical areas during the time judgment (p ≤ 0.025). CONCLUSION: The bromazepam decreases the precision of time estimation judgments and modulates the EEG alpha asymmetry, with greater left hemispheric dominance during time perception. Our findings suggest that bromazepam influences internal clock synchronization via the modulation of GABAergic receptors, strongly relating to attention, conscious perception, and behavioral performance.

Open monitoring meditation alters the EEG gamma coherence in experts meditators: The expert practice exhibit greater right intra-hemispheric functional coupling.

AIM: This study investigated the differences in frontoparietal EEG gamma coherence between expert meditators (EM) and naïve meditators (NM). MATERIAL AND METHODS: This is a cross-sectional study with a sample of twenty-one healthy adults divided under two groups (experts meditators vs. naive-meditators), with analyzing the intra-hemispheric coherence of frontoparietal gamma oscillations by electroencephalography during the study steps: EEG resting-state 1, during the open presence meditation practice, and EEG resting-state 2. RESULTS: The findings demonstrated greater frontoparietal EEG coherence in gamma for experts meditators in the Fp1-P3, F4-P4, F8-P4 electrode pairs during rest 1 and rest 2 (p ≤ 0.0083). In addition, we evidenced differences in the frontoparietal EEG coherence for expert meditators in F4-P4, F8-P4 during the meditation (p ≤ 0.0083). CONCLUSION: Our results can support evidence that the connectivity of the right frontoparietal network acts as a biomarker of the enhanced Open monitoring meditation training.

Non-immersive 3D virtual stimulus alter the time production task performance and increase the EEG theta power in dorsolateral prefrontal cortex.

AIM: The study investigated the cortical activity changes and time production task performance induced by changes in motion speed of a non-immersive 3D virtual stimulus. MATERIAL AND METHODS: Twenty-one individuals were participated in the crossover study with the visual-time reproduction task under three-speed conditions: original, slow and fast virtual stimulus. In addition, the electroencephalographic analysis of the theta band power in the dorsolateral prefrontal cortex was done simultaneously with time production task execution. RESULTS: The results demonstrated that in the slow speed condition, there is an increase in the error in the time production task after virtual reality (p < 0.05). There is also increased EEG theta power in the right dorsolateral prefrontal cortex in all speed conditions (p < 0.05). CONCLUSIONS: We propose that the modulations of speed of virtual stimulus may underlie the accumulation of temporal pulses, which could be responsible for changes in the performance of the production task of the time intervals and a substantial increase in right dorsolateral prefrontal cortex activity related to attention and memory, acting in cognitive domains of supraseconds.

Event-related potential (P300): the effects of levetiracetam in cognitive performance.

BACKGROUND: The current study is a reanalysis in the time domain of EEG data collection in healthy adults during an oddball paradigm using levetiracetam (LEV) vs. placebo acute administration. Specifically, the event-related potential (ERP) technique provides a tool for exploring the EEG responses to a specific event/stimulus. One of the ERP components widely studied is the P300 component, which is associated with the last stage of information processing and a general measurement of "cognitive efficiency." METHODS: The sample was composed of thirteen healthy right-handed individuals randomized to participate under two conditions: LEV and placebo. Electrophysiological measures were collected before and after drug intake. We explored the oddball paradigm, which is commonly used with healthy individuals to investigate the stages of information processing. RESULTS: The electrophysiological results showed a main effect of condition on P300 amplitude for the frontal (F3, Fz, F4), central (C3, Cz, C4), and parietal electrodes (P3, Pz, P4). The post hoc comparisons (Scheffé's test) demonstrated the significant differences between electrodes. Regarding P300 latency, all regions represented a main effect of condition. A P300 latency reduction was observed during LEV condition compared with placebo. CONCLUSION: Our study observed the ERP component-P300-through the variation of its amplitude and latency to evaluate a supposed higher CNS efficiency when participants were under the LEV effect. Our findings sustain this premise, mainly due to reducing in P300 latency for the LEV condition, supporting the neural efficiency hypothesis.

Time estimation exposure modifies cognitive aspects and cortical activity of attention deficit hyperactivity disorder adults.

AIM: This study investigated whether time-estimation task exposure influences the severity of Attention Deficit Hyperactivity Disorder (ADHD), as well as theta band activity in the dorsolateral prefrontal cortex and ventrolateral prefrontal cortex. MATERIAL AND METHODS: Twenty-two patients with ADHD participated in a crossover experiment with a visual time-estimation task under control conditions (without exposure to time estimation tasks) and experimental (thirty days exposure to time-estimation tasks) in association with electroencephalographic analysis of theta band. RESULTS: ADHD patients with thirty days of time-estimation task exposure presented a worse performance of the time-estimation task, as revealed by the measurements of the absolute error and relative error (p ≤ 0.05). However, our findings show the improvement of self-reported symptoms of attention, impulsivity, and emotional control in patients after the time-estimation task exposure (p = 0.0001). Moreover, the theta band oscillations in the right dorsolateral prefrontal cortex and in the ventrolateral prefrontal increased with thirty days of time-estimation task exposure (p ≤ 0.05). CONCLUSION: We propose that the decrease in EEG theta power may indicate an efficient accumulation of temporal pulses, which could be responsible for the improvement in the patient cognitive aspects as demonstrated by the current study. Time-estimation task improves ADHD cognitive symptoms, with a substantial increase in cortical areas activity related to attention and memory, suggesting its use as a tool for cognitive timing function management and non-invasive therapeutic aid in ADHD.

The role of low-frequency rTMS in the superior parietal cortex during time estimation.

The low-frequency repetitive transcranial magnetic stimulation (rTMS) application has been associated with changes in cognitive processes embedded during time perception tasks. Although several studies have investigated the influence of neuromodulation on time perception, the effect of the 1-Hz rTMS application on the superior parietal cortex is not clearly understood. This study analyzes the effect of the low-frequency rTMS on time estimation when applied in the parietal medial longitudinal fissure. For the proposed study, 20 subjects were randomly selected for a crossover study with two conditions (sham and 1 Hz). Our findings reveal that participant underestimate 1-s time interval and overestimate 4-s and 9-s time intervals after 1-Hz rTMS (p ≤ 0.05). We conclude that the 1-Hz rTMS in the parietal medial longitudinal fissure delays short interval and speed up long time intervals. This could be due to the effect of parietal inhibition on the attentional level and working memory functions during time estimation.

Methylphenidate modifies activity in the prefrontal and parietal cortex accelerating the time judgment.

Methylphenidate produces its effects via actions on cortical areas involved with attention and working memory, which have a direct role in time estimation judgment tasks. In particular, the prefrontal and parietal cortex has been the target of several studies to understand the effect of methylphenidate on executive functions and time interval perception. However, it has not yet been studied whether acute administration of methylphenidate influences performance in time estimation task and the changes in alpha band absolute power in the prefrontal and parietal cortex. The current study investigates the influence of the acute use of methylphenidate in both performance and judgment in the time estimation interpretation through the alpha band absolute power activity in the prefrontal and parietal cortex. This is a double-blind, crossover study with a sample of 32 subjects under control (placebo) and experimental (methylphenidate) conditions with absolute alpha band power analysis during a time estimation task. We observed that methylphenidate does not influence task performance (p > 0.05), but it increases the time interval underestimation by over 7 s (p < 0.001) with a concomitant decrease in absolute alpha band power in the ventrolateral prefrontal cortex and dorsolateral prefrontal cortex and parietal cortex (p < 0.001). Acute use of methylphenidate increases the time interval underestimation, consistent with reduced accuracy of the internal clock mechanisms. Furthermore, acute use of methylphenidate influences the absolute alpha band power over the dorsolateral prefrontal cortex, ventrolateral prefrontal cortex, and parietal cortex.

Low-frequency rTMS stimulation over superior parietal cortex medially improves time reproduction and increases the right dorsolateral prefrontal cortex predominance.

AIM OF THE STUDY: Previous studies have shown that several cortical regions are involved in temporal tasks in multiple timescales. However, the hemispheric predominance of the dorsolateral prefrontal cortex (DLPFC) during time reproduction after repetitive low-frequency transcranial magnetic stimulation (rTMS) is relatively unexplored. Here, we study the effects of 1 Hz rTMS and sham stimulation applied medially over the superior parietal cortex (SPC) on the DLPFC alpha and beta band asymmetry and on time reproduction. MATERIALS AND METHODS: For this purpose, we have combined rTMS with electroencephalography in 20 healthy subjects who performed the time reproduction task in two conditions (sham and 1 Hz). RESULTS: The worst performance was observed in sham and 1Hz conditions for longer time intervals (p < .05), with the 1Hz condition subjects sub-reproducing the time interval, closer to the target interval (p < .05). The right DLPFC hemispheric predominance was found in both conditions, but after low-frequency rTMS, the right hemisphere predominance increased in the 1Hz condition (p < .05). CONCLUSIONS: Results of this study suggest that rTMS applied over the SPC influences time interval interpretation and the DLPFC functions. Future studies would explore the effects of the rTMS application to other cortical areas, and study how it influences time interval interpretation.

Low-frequency rTMS in the superior parietal cortex affects the working memory in horizontal axis during the spatial task performance.

Spatial working memory has been extensively investigated with different tasks, treatments, and analysis tools. Several studies suggest that low frequency of the repetitive transcranial magnetic stimulation (rTMS) applied to the parietal cortex may influence spatial working memory (SWM). However, it is not yet known if after low-frequency rTMS applied to the superior parietal cortex, according to Pz electroencephalography (EEG) electrode, would change the orientation interpretation about the vertical and horizontal axes coordinates in an SWM task. The current study aims at filling this gap and obtains a better understanding of the low-frequency rTMS effect in SWM. In this crossover study, we select 20 healthy subjects in two conditions (control and 1-Hz rTMS). The subjects performed an SWM task with two random coordinates. Our results presented that low-frequency rTMS applied over the superior parietal cortex may influence the SWM to lead to a larger distance of axes interception point (p < 0.05). We conclude that low-frequency rTMS over the superior parietal cortex (SPC) changes the SWM performance, and it has more predominance in horizontal axis.

Proprioceptive neuromuscular facilitation increases alpha absolute power in the dorsolateral prefrontal cortex and superior parietal cortex.

The physiotherapist's clinical practice includes proprioceptive neuromuscular facilitation (PNF), which is a treatment concept that accelerates the response of neuromuscular mechanisms through spiral and diagonal movements. The adaptations that occur in the nervous system following PNF are still poorly described in the literature. Thus, this study had a goal to investigate the electrophysiological changes in the fronto-parietal circuit during PNF and movement in sagittal and diagonal patterns. This study included 30 female participants, who were divided into three groups (control, PNF, and flexion groups). Electroencephalogram measurements were determined before and after tasks were performed by each group. For the statistical analysis, a two-way ANOVA was performed for the factors group and time. Interactions between the two factors were investigated using a one-way ANOVA. A value of p < 0.004 was considered significant. The results showed an increase in alpha absolute power in the left dorsolateral prefrontal cortex and upper left parietal cortex of the PNF group, suggesting these areas work together to execute a motor action. The PNF group showed a greater alpha absolute power compared with the other groups, indicating a specific cortical demand for planning and attention, reinforcing its use for the rehabilitation of individuals.

Comparative analysis electroencephalographic of alpha, Beta and gamma bands of a healthy individual and one with hemiparesis.

[Purpose] The study analyzed the electroencephalographic (EEG) data of the central cortical areas, during execution of the motor gestures of feeding, activation of the system of mirror neurons, and imagery between a right hemiparetic volunteer (RHV) and a healthy volunteer (HV). [Subjects and Methods] The volunteers' EEG data were recorded with their eyes open for 4 minutes while they performed five experimental tasks. [Results] The alpha band, absolute power value of HV was lower than that of RHV. In the beta band, during the practice condition, there was an increase in the magnitude of the absolute power value of HV at T3, possibly because T3 is representative of secondary motor areas that work with cortical neurons related to planning and organizing sequence of movements performed by the hands. The gamma band is related to the state of preparation for movement and memory. The results of this study indicate that there was increased activation of the gamma frequency band of HV. [Conclusion] The findings of this study have revealed the changes in pattern characteristics of each band which may be associated with the brain injury of the hemiparetic patient.

Curve-Modelling and Machine Learning for a Better COPD Diagnosis.

Background: Development of new tools in artificial intelligence has an outstanding performance in the recognition of multidimensional patterns, which is why they have proven to be useful in the diagnosis of Chronic Obstructive Pulmonary Disease (COPD). Methods: This was an observational analytical single-centre study in patients with spirometry performed in outpatient medical care. The segment that goes from the peak expiratory flow to the forced vital capacity was modelled with quadratic polynomials, the coefficients obtained were used to train and test neural networks in the task of classifying patients with COPD. Results: A total of 695 patient records were included in the analysis. The COPD group was significantly older than the No COPD group. The pre-bronchodilator (Pre BD) and post-bronchodilator (Post BD) spirometric curves were modelled with a quadratic polynomial, and the coefficients obtained were used to feed three neural networks (Pre BD, Post BD and all coefficients). The best neural network was the one that used the post-bronchodilator coefficients, which has an input layer of 3 neurons and three hidden layers with sigmoid activation function and two neurons in the output layer with softmax activation function. This system had an accuracy of 92.9% accuracy, a sensitivity of 88.2% and a specificity of 94.3% when assessed using expert judgment as the reference test. It also showed better performance than the current gold standard, especially in specificity and negative predictive value. Conclusion: Artificial Neural Networks fed with coefficients obtained from quadratic and cubic polynomials have interesting potential of emulating the clinical diagnostic process and can become an important aid in primary care to help diagnose COPD in an early stage.

Decrease in reaction time for volleyball athletes during saccadic eye movement task: A preliminary study with evoked potentials.

AIM: This preliminary study investigated the differences in event-related potential and reaction time under two groups (athletes vs. non-athletes). MATERIAL AND METHODS: The P300 was analyzed for Fz, Cz, and Pz electrodes in thirty-one healthy volunteers divided into two groups (volleyball athletes and non-athletes). In addition, the participants performed a saccadic eye movement task to measure reaction time. RESULTS: The EEG analysis showed that the athletes, in comparison to the no-athletes, have differences in the P300 in the frontal area (p = 0.021). In relation to reaction time, the results show lower reaction time for athletes (p = 0.001). CONCLUSIONS: The volleyball athletes may present a greater allocation of attention during the execution of the inhibition task, since they have a lower reaction time for responses when compared to non-athletes.

Changes in Cortical Activity During Real and Imagined Movements: an ERP Study.

This study aims to compare the topographic distribution of cortical activation between real and imagined movement through event-related potential (ERP). We are specifically interested in identifying, the topographic distribution of activated areas, the intensity of activated areas, and the temporal occurrence of these activations on preparation and motor response phases. Twelve healthy and right handed subjects were instructed to perform a task under real and imagery conditions. The task was performed simultaneously to electroencephalographic (EEG) recording. When compared the conditions, we found a statistically significant difference in favor of real condition revealed by performing an unpaired t-test with multiple corrections of Bonferroni, demonstrating negative activity on electrode C3 and positive activity on the electrode C4 only in motor response phase. These findings revealed similar functional connections established during real and imagery conditions, suggesting that there are common neural substrate and similar properties of functional integration shared by conditions.

Differences between hemispheres and in saccade latency regarding volleyball athletes and non-athletes during saccadic eye movements: an analysis using EEG.

BACKGROUND: The saccadic eye movement is responsible for providing focus to a visual object of interest to the retina. In sports like volleyball, identifying relevant targets quickly is essential to a masterful performance. The training improves cortical regions underlying saccadic action, enabling more automated processing in athletes. OBJECTIVE: We investigated changes in the latency during the saccadic eye movement and the absolute theta power on the frontal and prefrontal cortices during the execution of the saccadic eye movement task in volleyball athletes and non-athletes. We hypothesized that the saccade latency and theta power would be lower due to training and perceptual-cognitive enhancement in volleyball players. METHODS: We recruited 30 healthy volunteers: 15 volleyball athletes (11 men and 4 women; mean age: 15.08 ± 1.06 years) and 15 non-athletes (5 men and 10 women; mean age: 18.00 ± 1.46 years). All tasks were performed simultaneously with electroencephalography signal recording. RESULTS: The latency of the saccadic eye movement presented a significant difference between the groups; a shorter time was observed among the athletes, associated with the players' superiority in terms of attention level. During the experiment, the athletes observed a decrease in absolute theta power compared to non-athletes on the electrodes of each frontal and prefrontal area. CONCLUSION: In the present study, we observed the behavior of reaction time and absolute theta power in athletes and non-athletes during a saccadic movement task. Our findings corroborate the premise of cognitive improvement, mainly due to the reduction of saccadic latency and lower beta power, validating the neural efficiency hypothesis.

ANTECEDENTES: O movimento ocular sacádico é responsável por dar foco a um objeto visual de interesse para a retina. Em esportes como o vôlei, identificar alvos relevantes o mais rápido possível é essencial para se ter um desempenho magistral. O treinamento melhora as regiões corticais subjacentes à ação sacádica, e permite um processamento mais automatizado em atletas. OBJETIVO: Investigamos as mudanças na latência durante o movimento ocular sacádico e a potência teta absoluta nos córtices frontal e pré-frontal durante a execução da tarefa de movimento ocular sacádico em atletas e não atletas de voleibol. Nossa hipótese é a de que a latência sacádica e a potência teta seriam menores em atletas devido ao treinamento e ao aprimoramento perceptivo-cognitivo em jogadores de voleibol. MéTODOS: Ao todo, 30 voluntários saudáveis foram recrutados para este estudo: 15 atletas de voleibol (11 homens e 4 mulheres; idade média: 15,08 ± 1,06 anos) e 15 não atletas (5 homens e 10 mulheres; idade média: 18,00 ± 1,46 anos). Todas as tarefas foram realizadas simultaneamente com o registro do sinal eletroencefalográfico. RESULTADOS: O resultado da latência do movimento ocular sacádico apresentou diferença significativa entre os grupos, sendo observado menor tempo entre os atletas, associado à superioridade dos jogadores em termos de nível de atenção. Durante o experimento, nos eletrodos de cada área frontal e pré-frontal, observou-se uma diminuição na potência teta absoluta nos atletas em comparação aos não atletas. CONCLUSãO: Neste estudo, observou-se o comportamento do tempo de reação e da potência teta absoluta em atletas e não atletas durante uma tarefa de movimento sacádico. Nossos achados corroboram a premissa de melhora cognitiva, principalmente pela redução da latência sacádica e menor potência beta, o que valida a hipótese de eficiência neural.

Dietary habits and inadequate control of blood pressure in hypertensive adults assisted by a Brazilian Family Doctor Program.

OBJECTIVE: To estimate dietary habits and other factors associated with inadequate blood pressure (BP) control in hypertensive patients adherent to antihypertensive drug treatment assisted by a Brazilian Family Doctor Program (FDP). DESIGN: A cross-sectional study. SETTING: FDP units, Niterói, Rio de Janeiro, Brazil. SUBJECTS: We included data from both male and female participants aged ≥20 years. Participants completed a standardized questionnaire containing questions related to demographics, socio-economic factors, comorbidities and lifestyle, as well as a validated FFQ and eleven additional qualitative questions to investigate dietary habits. Food items were divided into sixteen groups. Medical consultations were performed, BP measurements were taken, blood and urine samples were assessed and anthropometric and nutritional status was evaluated. RESULTS: Individuals with inadequate BP control presented higher BMI values (prevalence ratio (PR) = 1·027, 95% CI 1·009, 1·045) and also consumed more meat (PR = 1·091, 95% CI 1·022, 1·165), which are potentially modifiable variables. Higher levels of serum creatinine (PR = 1·894, 95% CI 1·241, 2·892) were also associated with inadequate BP control, as were skin colour (white). After inclusion of the Na excretion index, which is an indirect measure of salt intake, a slight decrease was observed in the PR for meat, which resulted in loss of statistical significance. CONCLUSIONS: The results indicate that salt consumption, skin colour, BMI and serum creatinine are associated with inadequate BP control.

The effect of acute effort on EEG in healthy young and elderly subjects.

The effects of physical exercise on mental health have been extensively investigated, mainly in older people. Recent studies have looked into the acute effect of exercise on the brain using standardized low-resolution brain electromagnetic tomography (sLORETA). We assessed EEG power and mood changes after 20 min of aerobic exercise in elderly (N = 10) and young (N = 19) healthy individuals. Both groups showed improvement in total mood disturbance (TMD) post exercise (young: P = 0.03; elderly: P = 0.02). Only the young group showed significant improvement in anger (P = 0.05) and vigor (P = 0.006). Comparison pre versus post-exercise for each group separately revealed significant changes in the young group (an increase in alpha, beta-1 and beta-2 activity in Brodmann areas 24, 33 and 23, respectively). However, the elderly group did not show significant changes. An inverse correlation was found between alpha asymmetry and STAI (rs = -0.50; P = 0.029) in the young group. On the other hand, a significant correlation between beta-1 activity and TMD was observed in the elderly group (rs = 0.67; P = 0.045). We conclude that acute exercise can have distinct effects on brain activity and mood variables in young individuals when compared with elderly adults. However, additional studies are necessary to further investigate the role of exercise intensity in these results.

Development of a Classical Conditioning Task for Humans Examining Phasic Heart Rate Responses to Signaled Appetitive Stimuli: A Pilot Study.

Cardiac responses to appetitive stimuli have been studied as indices of motivational states and attentional processes, the former being associated with cardiac acceleration and latter deceleration. Very few studies have examined heart rate changes in appetitive classical conditioning in humans. The current study describes the development and pilot testing of a classical conditioning task to assess cardiac responses to appetitive stimuli and cues that reliably precede them. Data from 18 adults were examined. They were shown initially neutral visual stimuli (putative CS) on a computer screen followed by pictures of high-caloric food (US). Phasic cardiac deceleration to food images was observed, consistent with an orienting response to motivationally significant stimuli. Similar responses were observed to non-appetitive stimuli when they were preceded by the cue associated with the food images, suggesting that attentional processes were engaged by conditioned stimuli. These autonomic changes provide significant information about classical conditioning effects in humans.

Repetitive transcranial magnetic stimulation changes cognitive/motor tasks performance: An absolute alpha and beta power study.

The voluntary movement demands integration between cognitive and motor functions. During the initial stages of motor learning until mastery of a new motor task, and during a demanding task that is not automatic, cognitive and motor functions can be perceived as independent from each other. Areas used for actually performing motor tasks are essentially the same used by Motor Imagery (MI). The main objective of this study was to investigate inhibition effects on cognitive functions of motor skills induced by low-frequency (1 Hz) Repetitive Transcranial Magnetic Stimulation (rTMS) at the sensory-motor integration site (Cz). In particular, the goal was to examine absolute alpha and beta power changes on frontal regions during Execution, Action observation, and Motor Imagery of finger movement tasks. Eleven healthy, right-handed volunteers of both sexes (5 males, 6 females; mean age 28 ± 5 years), with no history of psychiatric or neurological disorders, participated in the experiment. The execution task consisted of the subject flexing and extending the index finger. The action observation task involved watching a video of the same movement. The motor imagery task was imagining the flexion and extension of the index finger movement. After performing the tasks randomly, subjects were submitted to 15 min of low-frequency rTMS and performed the tasks again. All tasks were executed simultaneously with EEG signals recording. Our results demonstrated a significant interaction between rTMS and the three tasks in almost all analyzed regions showing that rTMS can affect the frontal region regarding Execution, Action observation, and Motor Imagery tasks.

Methylphenidate decreases the EEG mu power in the right primary motor cortex in healthy adults during motor imagery and execution.

This study investigated the effects of dopaminergic drugs on the EEG mu power during motor imagery, action observation, and execution. This is a double-blind, crossover study with a sample of 15 healthy adults under placebo vs. methylphenidate vs. risperidone conditions during motor imagery, action observation, and execution tasks. The participants had drug dosage adjustment based on body weight/dose (mg/kg). We also analyzed the mu band power by electroencephalography during the study steps. The main result is the interaction between the condition and task factors for the C3 and C4 electrodes, with decreasing EEG mu power in the methylphenidate when compared to risperidone (p ≤ 0.0083). Our results can indicate that the methylphenidate decreases the neurophysiological activity in the central cortical regions during the perceptual experience of tasks with or without body movement.