Novel measures of Morris water maze performance that use vector field maps to assess accuracy, uncertainty, and intention of navigational searches.

Most commonly used behavioral measures for testing learning and memory in the Morris water maze (MWM) involve comparisons of an animal's residence time in different quadrants of the pool. Such measures are limited in their ability to test different aspects of the animal's performance. Here, we describe novel measures of performance in the MWM that use vector fields to capture the motion of mice as well as their search pattern in the maze. Using these vector fields, we develop quantitative measures of performance that are intuitive and more sensitive than classical measures. First, we describe search patterns in terms of vector field properties and use these properties to define three metrics of spatial memory namely Spatial Accuracy, Uncertainty and, Intensity of Search. We demonstrate the usefulness of these measures using four different data sets including comparisons between different strains of mice, an analysis of two mouse models of Noonan syndrome (NS; Ptpn11 D61G and Ptpn11 N308D/+), and a study of goal reversal training. Importantly, besides highlighting novel aspects of performance in this widely used spatial task, our measures were able to uncover previously undetected differences, including in an animal model of NS, which we rescued with the mitogen activated protein kinase kinase (MEK) inhibitor SL327. Thus, our results show that our approach breaks down performance in the MWM into sensitive measurable independent components that highlight differences in spatial learning and memory in the MWM that were undetected by conventional measures.

A new paradigm for investigating temporal order memory shows higher order associations are present in recent but not in remote retrieval.

Memory of a sequence of distinct events requires encoding the temporal order as well as the intervals that separates these events. In this study, using order-place association task where the animal learns to associate the location of the food pellet to the order of entry into the event arena, we probe the nature of temporal order memory in mice. In our task, individual trials become distinct events, as the animal is trained to form a unique association between entry order and a correct location. The inter-trial intervals (> 30 min) are chosen deliberately to minimize the inputs from working memory. We develop this paradigm initially using four order-place associates and later extend it to five paired associates. Our results show that animals not only acquire these explicit (entry order to place) associations but also higher order associations that can only be inferred implicitly (temporal relation between the events) from the temporal order of these events. As an indicator of such higher order learning during the probe trial, the mice exhibit predominantly prospective errors that decline proportionally with temporal distance. On the other hand, prior to acquiring the sequence, the retrospective errors are dominant. In addition, we also tested the nature of such acquisitions when temporal order CS is presented along with flavored pellet as a compound stimulus comprising of order and flavor both simultaneously being paired with location. Results from these experiments indicate that the animal learns both order-place and flavor-place associations. Comparing with pure order-place training, we find that the additional flavor stimulus in a compound training paradigm did not interfere with the ability of the animals to acquire the order-place associations. When tested remotely, pure order-place associations could be retrieved only after a reminder training. Further higher order associations representing the temporal relationship between the events is markedly absent in the remote time.

Saturation Dynamics Measures Absolute Cross Section and Generates Contrast within a Neuron.

The intensity required to optically saturate a chromophore is a molecular property that is determined by its absorption cross section (σ) and the excited state lifetime. We present an analytical description of such a system and show that fluorescence around the onset of saturation is characterized by product of absorption cross section and lifetime. Using this approach we formulate a generalized method for measuring the multiphoton cross section of fluorophores and use it to obtain the absolute three-photon cross-section spectra of tryptophan. We find that the tryptophan three-photon cross section ranges from 0.28 S.I. units (m(6)s(2)photon(-2)) at 870 nm to 20 S.I. units at 740 nm. Further, we show that the product of molecular rate of excitation and de-excitation, denoted as ß, serves as a vital contrasting agent for imaging local environment. Our contrast parameter, ß, is related to fraction of the population present in the excited state and is independent of the fluorophore concentration. We show that ß-imaging can be carried out in a regular two-photon microscope setup through a series of intensity scans. Using enhanced green fluorescent protein (EGFP) fluorescence from the brain slices of Thy-1 EGFP transgenic mice, we show that there is an inherent, concentration independent, variation in contrast across the soma and the dendrite.

Mortality risk among tobacco users in a rural population of southern India: evidence from a cohort study.

BACKGROUND: Tobacco has long been identified as one of the most hazardous risk factors detrimental to health. To plan and implement anti-tobacco activities in any community, it is necessary to understand the risk it poses in that setting. We assessed the risk of mortality associated with tobacco use in a rural community of Kerala. METHODS: This cohort study (PROLIFE) was done in Varkkala, a rural development block of Thiruvananthapuram district of Kerala. Adults aged 20 years and above were included. Age-adjusted mortality rates were computed for both users and non-users of tobacco. The risk of mortality was plotted using Kaplan-Meier curves. Cox regression was used to compute the age-adjusted hazard ratio of mortality among tobacco users. RESULTS: More than one-fourth of the study population used tobacco. The age-adjusted mortality rates were higher among tobacco users. The major causes of death among both users and non-users of tobacco were similar. The incidence proportion of death among all causes of death was higher for tobacco users. The hazard of mortality was significantly more among tobacco users, with the age-adjusted hazard ratio being 1.225 (1.140-1.315). CONCLUSION: The mortality risk due to tobacco use is high irrespective of the cause of death.

Crystal structure of (E)-N'-(4-chloro-benzyl-idene)-4-methyl-benzene-sulfono-hydrazide: a hexa-gonal polymorph.

The title compound, C14H13ClN2O2S, crystallized in the enanti-omorphic defining hexa-gonal space group P61 [Flack parameter = -0.02 (7)]. The partially hydrated form of the same compound, crystallizing in the triclinic space group P-1, has been reported previously [Kia et al. (2009b). Acta Cryst. E65, o1119], as has the crystal structure of the bromo derivative, also crystallizing in the space group P-1 [Kia et al. (2009a). Acta Cryst. E65, o821]. The title mol-ecule is non-planar with the planes of the benzene rings being inclined to one another by 76.62 (13)°, and has an E conformation about the C=N bond. In the crystal, mol-ecules are linked via N-H⋯O hydrogen bonds forming 61 helical chains running along [001]. The chains are linked via C-H⋯O hydrogen bonds, C-H⋯π inter-actions and short Cl⋯O [3.015 (3) Å] inter-actions, forming a three-dimensional structure.

Hybrid manifold smoothing and label propagation technique for Kannada handwritten character recognition.

Handwritten character recognition is one of the classical problems in the field of image classification. Supervised learning techniques using deep learning models are highly effective in their application to handwritten character recognition. However, they require a large dataset of labeled samples to achieve good accuracies. Recent supervised learning techniques for Kannada handwritten character recognition have state of the art accuracy and perform well over a large range of input variations. In this work, a framework is proposed for the Kannada language that incorporates techniques from semi-supervised learning. The framework uses features extracted from a convolutional neural network backbone and uses regularization to improve the trained features and label propagation to classify previously unseen characters. The episodic learning framework is used to validate the framework. Twenty-four classes are used for pre-training, 12 classes are used for testing and 11 classes are used for validation. Fine-tuning is tested using one example per unseen class and five examples per unseen class. Through experimentation the components of the network are implemented in Python using the Pytorch library. It is shown that the accuracy obtained 99.13% make this framework competitive with the currently available supervised learning counterparts, despite the large reduction in the number of labeled samples available for the novel classes.

Familial Gigantiform Cementoma: Life-Saving Total Midface Resection and Reconstruction Using Virtual Surgical Planning and 3D Printed Patient-Specific Implant-A Clinical Study.

Background: Familial gigantiform cementoma (FGC) is a rare benign fibrocemento-osseous lesion of the jaw characterized by well-circumscribed, extensive, mixed radiolucent-radiopaque masses in the mandible and the maxilla that can cause severe facial deformity. This condition is extremely rare with less than 40 cases reported in the literature. Purpose: The purpose of the paper is to highlight the importance of virtual surgical planning and patient-specific implant in the treatment of a complex lesion and reconstruction of the facial skeleton. The clinical presentations, and diagnostic challenges encountered when managing the lesion have been discussed in this article with emphasis on the treatment plan. Method/Surgical plan: The sequence of treatment planned was resection of the lesion and immediate reconstruction with a patient-specific implant to improve the patient's quality of life. The management of FGC was a challenging one keeping in mind the rapid expansion of the lesion, widespread involvement of the jaws, and needs of the pediatric patient. Conclusion: Virtual surgical planning (VSP) along with 3D printed implant was instrumental in reconstructing the facial form of the child where the maxilla was completely resected and rehabilitation provided support to the vital structures of the face.

Unicystic ameloblastoma - A case series.

Ameloblastoma is a benign, locally aggressive neoplasm that constitutes about 1-3% of the tumors of the jaw. Wide surgical excision with adequate safe margin is the most common treatment of choice. The study aimed to manage cases with unicystic ameloblastoma while preserving the continuity of the mandible (without resection). This article presents a series of cases ranging from 18 to 40 years old patients of both sexes with unicystic ameloblastoma, especially in the mandible showing more male predilection than female. All the cases presented in this article were treated by enucleation and curettage. None of the patients presented post-operative paresthesia. None of the cases went in for resection. Post-operative recovery was uneventful in all the patients. All the patients were followed up for a period of 3.5-5 years. None of the cases reported recurrence at the date of publication.

A family of familial gigantiform cementoma: clinical study.

Background: Familial gigantiform cementoma is an autosomal dominant fibro-cement osseous lesion that causes massive expansion of facial skeleton. Patients with such massive lesions have a compromised quality of life. The main goal of treating such patients is to restore and preserve the jaw as far as possible which would enhance their quality of life. Purpose: This study was conducted to identify the occurrence of gigantiform cementoma which had affected three generations of a family and also to focus on documentation of the clinical course and management. Method: Patients (one family-mother, grandmother, aunt and grandson) who had visited the Department of Oral and Maxillofacial Surgery, Tamilnadu Government Dental College and Hospital, Chennai, over a period for their swelling in the maxillofacial region were clinically and radiographically evaluated and histopathologically diagnosed as familial gigantiform cementoma; later, they were surgically managed. Result: It is very rare to document three generations of this disease which had shown varied clinical presentation (asymptomatic slow growth, arrested growth and one case of aggressive growth). Management of these cases varied from observation to aggressive resection. Conclusion: Gigantiform cementoma follows an autosomal dominant pattern of inheritance with variable phenotypic expression without gender predilection. These cases require regular observation and intervention if necessary.

Effect of silver diamine fluoride, potassium nitrate, and glutaraldehyde in reducing the post vital tooth preparation hypersensitivity: A randomized controlled trial.

Aim: Hypersensitivity is the most common clinical problem which is encountered by most of dental patients undergoing a vital tooth preparation for a fixed crown prosthesis. The aim of this study is to evaluate the effect of silver diamine fluoride, potassium nitrate, and glutaraldehyde in reducing dentinal hypersensitivity following vital tooth preparation. Settings and Design: This study is a randomized control trial performed on 119 teeth of 68 patients who are in need of fixed prosthesis treatment. Materials and Methods: After a thorough clinical examination, patients were allocated into any of the randomly assigned four groups (Control, silver diamine fluoride, potassium nitrate, and glutaraldehyde) and the level of hypersensitivity was measured by blasting air on the surface of tooth at five different intervals (before preparation, after preparation, after application of desensitizers, before cementation and after a follow up period of about 30 days) and is graded using Schiff 's cumulative hypersensitivity index. Statistical Analysis Used: Kruskal wallis test is used to compare the rate of sensitivity between the 4 groups. Friedman and Wilcoxon test is used to compare the rate of sensitivity at 5 different intervals. Results: All the desensitizers used in this study reduced the level of hypersensitivity. Among which, silver diamine fluoride was found to be more effective after application, before cementation, and after a follow up period of about 30 days followed by GLUMA and potassium nitrate. Conclusion: The results of this study suggest that silver diamine fluoride was found to be more effective after application, before cementation of the definitive prosthesis and after a follow up period of 30 days.

In vivo imaging of immediate early gene expression dynamics segregates neuronal ensemble of memories of dual events.

Identification of neurons undergoing plasticity in response to external stimuli is one of the pertinent problems in neuroscience. Immediate early genes (IEGs) are widely used as a marker for neuronal plasticity. Here, we model the dynamics of IEG expression as a consecutive, irreversible first-order reaction with a limiting substrate. First, we develop an analytical framework to show that such a model, together with two-photon in vivo imaging of IEG expression, can be used to identify distinct neuronal subsets representing multiple memories. Using the above combination, we show that the expression kinetics, rather than intensity threshold, can be used to identify neuronal ensembles responding to the presentation of two events in vivo. The analytical expression allowed us to segregate the neurons based on their temporal response to one specific behavioural event, thereby improving the ability to detect plasticity related neurons. We image the retrosplenial cortex (RSc) of cfos-GFP transgenic mice to follow the dynamics of cellular changes resulting from contextual fear conditioning behaviour, enabling us to establish a representation of context in RSc at the cellular scale following memory acquisition. Thus, we obtain a general method that distinguishes neurons that took part in multiple temporally separated events by measuring fluorescence of individual neurons in live mice.

Single-vesicle imaging reveals that synaptic vesicle exocytosis and endocytosis are coupled by a single stochastic mode.

The nature of synaptic vesicle recycling at nerve terminals has been a subject of considerable debate for >35 years. Here, we report the use of an optical strategy that allows the exocytosis and retrieval of synaptic components to be tracked in real time at single-molecule sensitivity in living nerve terminals. This approach has allowed us to examine the recycling of synaptic vesicles in response to single action potentials. Our results show that, after exocytosis, individual synaptic vesicles are retrieved by a stochastic process with an exponential distribution of delay times, with a mean time of approximately 14 s. We propose that evidence for fast endocytosis, such as that proposed to support the presence of kiss-and-run, is likely explained by the stochastic nature of a slower process.

White-Eyed Blowout Fracture.

White-eyed blowout fracture is often found in pure orbital floor blowout fracture among pediatric patients. Unlike common orbital blowout fractures with apparent clinical signs, the diagnosis of white-eyed orbital blowout fractures is difficult because of minimal soft-tissue signs. This report describes an early missed-out diagnosis of a white-eyed blowout fracture in a 7-year-old child, due to negligible soft-tissue manifestation.

Six-hole versus Four-hole Miniplates in Isolated, Unilateral Angle Fracture of the Mandible.

AIM: The aim of the study was to find the suitable situation for the fixation of "six" hole miniplates in open reduction and internal fixation of displaced and nondisplaced unilateral angle of the mandible fractures. SUBJECTS AND METHODS: Displaced, nondisplaced, simple, compound (linear and noncomminuted) isolated unilateral angle fractures with or without occlusal derangement were included in this study. STATISTICAL ANALYSIS USED: Chi-square test was used for the statistical analysis. RESULTS: The parameters assessed preoperatively and postoperatively were mouth opening, occlusion, neurosensory deficit such as paresthesia, intraoperative time, stability of fragments, and access to the retromolar trigone which showed that the six-hole titanium miniplate was clinically useful in special clinical situations when compared to four-hole titanium miniplate. CONCLUSION: To conclude, six-hole titanium miniplate was clinically useful when compared to four-hole titanium miniplate in the following special clinical situations such as bone loss following extraction of third molar, no posterior occlusion and instability of fracture moderately displaced fracture needs more stability.

Calcium control of endocytic capacity at a CNS synapse.

The ability to recycle synaptic vesicles is a crucial property of nerve terminals that allows maintenance of synaptic transmission. Using high-sensitivity optical approaches at hippocampal nerve terminals in dissociated neurons in culture, we show that modulation of endocytosis can be achieved by expansion of the endocytic capacity. Our experiments indicate that the endocytic capacity, the maximum number of synaptic vesicles that can be internalized in parallel at individual synapses, is tightly controlled by intracellular calcium levels. Increasing levels of intracellular calcium, which occurs as firing frequency increases, significantly increases the endocytic capacity. At physiological temperature after 30 Hz firing, these synapses are capable of endocytosing at least approximately 28 vesicles in parallel, each with a time constant of approximately 6 s. This calcium-dependent control of endocytic capacity reveals a potentially useful adaptive response to high-frequency activity to increase endocytic rates under conditions of vesicle pool depletion.

Intermolecular association provides specific optical and NMR signatures for serotonin at intravesicular concentrations.

Neurotransmitter vesicles contain biomolecules at extraordinarily high concentrations (hundreds of millimoles/liter). Such concentrations can drive intermolecular associations, which may affect vesicular osmolarity and neuronal signaling. Here we investigate whether aqueous serotonin (a monoamine neurotransmitter) forms oligomers at intravesicular concentrations and whether these oligomers have specific spectroscopic signatures that can potentially be used for monitoring neuronal storage and release. We report that, as serotonin concentration is increased from 60 microM to 600 mM, the normalized fluorescence spectrum of serotonin displays a growing long-wavelength tail, with an isoemissive point at 376 nm. The fluorescence decay is monoexponential with a lifetime of 4 ns at low concentrations but is multiexponential with an average lifetime of 0.41 ns at 600 mM. A 600 mM serotonin solution has 30% less osmolarity than expected for monomeric serotonin, indicating oligomer formation. The proton NMR chemical shifts move upfield by as much as 0.3 ppm at 600 mM compared to those at 10 mM, indicating a stacking of the serotonin indole moieties. However, no intermolecular crosspeak is evident in the two-dimensional NMR rotating frame Overhauser effect spectroscopy spectrum even at 600 mM, suggesting that oligomeric structures are possibly weakly coupled. The appearance of a single peak for each proton suggests that the rate of interconversion between the monomeric and the oligomeric structures is faster than 240 Hz. A stopped-flow kinetic experiment also confirms that the rate of dissociation is faster than 100 ms. We conclude that serotonin forms oligomers at intravesicular concentrations but becomes monomeric quickly on dilution. NMR signatures of the oligomers provide potential contrast agents for monitoring the activity of serotonergic neurons in vivo.

Three-photon microscopy shows that somatic release can be a quantitatively significant component of serotonergic neurotransmission in the mammalian brain.

Recent experiments on monoaminergic neurons have shown that neurotransmission can originate from somatic release. However, little is known about the quantity of monoamine available to be released through this extrasynaptic pathway or about the intracellular dynamics that mediate such release. Using three-photon microscopy, we directly imaged serotonin autofluorescence and investigated the total serotonin content, release competence, and release kinetics of somatic serotonergic vesicles in the dorsal raphe neurons of the rat. We found that the somata of primary cultured neurons contain a large number of serotonin-filled vesicles arranged in a perinuclear fashion. A similar distribution is also observed in fresh tissue slice preparations obtained from the rat dorsal raphe. We estimate that the soma of a cultured neuron on an average contains about 9 fmoles of serotonin in about 450 vesicles (or vesicle clusters) of < or =370 nm average diameter. A substantial fraction (>30%) of this serotonin is released with a time scale of several minutes by K(+)-induced depolarization or by para-chloroamphetamine treatment. The amount of releasable serotonin stored in the somatic vesicles is comparable to the total serotonin content of all the synaptic vesicles in a raphe neuron, indicating that somatic release can potentially play a major role in serotonergic neurotransmission in the mammalian brain.

Serotonin in pre-implantation mouse embryos is localized to the mitochondria and can modulate mitochondrial potential.

Serotonin is reported to be present in early embryos of many species and plays an important role in early patterning. Since it is a fluorophore, it can be directly visualized using fluorescence microscopy. Here, we use three-photon microscopy to image serotonin in live pre-implantation mouse embryos. We find that it is present as puncta averaging 1.3 square microns and in concentrations as high as 442 mM. The observed serotonin puncta were found to co-localize with mitochondria. Live embryos pre-incubated with serotonin showed a higher mitochondrial potential, indicating that it can modulate mitochondrial potential. Pre-implantation mouse embryos were also examined at various developmental stages for the presence of transcripts of the peripheral and neuronal forms of tryptophan hydroxylase (Tph1 and Tph2 respectively) and the classical serotonin transporter (Slc6a4). Transcripts of Tph2 were seen in oocytes and in two-cell stages, whereas transcripts of Tph1 were not detected at any stage. Transcripts of the transporter, Slc6a4, were present in all pre-implantation stages investigated. These results suggest that serotonin in embryos can arise from a combination of synthesis and uptake from the surrounding milieu.

Protein aggregation probed by two-photon fluorescence correlation spectroscopy of native tryptophan.

Fluorescence correlation spectroscopy (FCS) has proven to be a powerful tool for the study of a range of biophysical problems including protein aggregation. However, the requirement of fluorescent labeling has been a major drawback of this approach. Here we show that the intrinsic tryptophan fluorescence, excited via a two-photon mechanism, can be effectively used to study the aggregation of tryptophan containing proteins by FCS. This method can also yield the tryptophan fluorescence lifetime in parallel, which provides a complementary parameter to understand the aggregation process. We demonstrate that the formation of soluble aggregates of barstar at pH 3.5 shows clear signatures both in the two-photon tryptophan FCS data and in the tryptophan lifetime analysis. The ability to probe the soluble aggregates of unmodified proteins is significant, given the major role played by this species in amyloid toxicity.