Melanocortin-4 receptor signaling is not required for short-term weight loss after sleeve gastrectomy in pediatric patients.

Homozygous or compound heterozygous melanocortin-4 receptor (MC4R) mutations are rare with fewer than 10 patients described in current literature. Here we report the short- and long-term outcomes for four children ages 4.5-14 who are homozygous for loss-of-function mutations in the MC4R and underwent laparoscopic sleeve gastrectomy. All four patients experienced significant weight loss and improvement in, or resolution of, their comorbidities in the short term. One patient, however, has had significant weight regain in the long term. We conclude that MC4R signaling is not required for short-term weight loss after laparoscopic sleeve gastrectomy in children. Behavior modification may be more important for long-term weight maintenance, but patients with homozygous MC4R deficiency should not be excluded from consideration for sleeve gastrectomy. However, as at least one copy of functional MC4R is necessary and sufficient to induce long-term postoperative weight loss benefits, patients with complete loss of MC4R functionality might be less likely to exhibit the same benefits resulting from bariatric surgery.

Differentiation of human dental stem cells reveals a role for microRNA-218.

BACKGROUND: Regeneration of lost periodontium is the ultimate goal of periodontal therapy. Advances in tissue engineering have demonstrated the multilineage potential and plasticity of adult stem cells located in periodontal apparatus. However, it remains unclear how epigenetic mechanisms controlling signals determine tissue specification and cell lineage decisions. To date, no data are available on micro-RNA (miRNA) activity behind human-derived dental stem cells (DSCs). MATERIAL AND METHODS: In this study, we isolated periodontal ligament stem cells, dental pulp stem cells and gingival stem cells from extracted third molars; human bone marrow stem cells were used as a positive control. The expression of OCT4A and NANOG was confirmed in these undifferentiated cells. All cells were cultured under osteogenic inductive conditions and RUNX2 expression was analyzed as a marker of mineralized tissue differentiation. The miRNA expression profile was obtained at baseline and after osteogenic induction in all cell types. RESULTS: The expression of RUNX2 demonstrated successful osteogenic induction of all cell types, which was confirmed by alizarin red stain. The analysis of 765 miRNAs demonstrated a shift in miRNA expression that occurred in all four stem cell types, including a decrease in hsa-mir-218 across all differentiated cell populations. Hsa-mir-218 targets RUNX2 and decreases RUNX2 expression in undifferentiated human DSCs. DSC mineralized tissue type differentiation is associated with a decrease in hsa-mir-218 expression. CONCLUSION: These data reveal a miRNA-regulated pathway for the differentiation of human DSCs and a select network of human miRNAs that control DSC osteogenic differentiation.

Mycobacterium tuberculosis Toxin CpnT Is an ESX-5 Substrate and Requires Three Type VII Secretion Systems for Intracellular Secretion.

CpnT, a NAD+ glycohydrolase, is the only known toxin that is secreted by Mycobacterium tuberculosis CpnT is composed of two domains; the C-terminal domain is the toxin, whereas the N-terminal domain is required for secretion. CpnT shows characteristics of type VII secretion (T7S) substrates, including a predicted helix-turn-helix domain followed by a secretion motif (YxxxE). Disruption of this motif indeed abolished CpnT secretion. By analyzing different mutants, we established that CpnT is specifically secreted by the ESX-5 system in Mycobacterium marinum under axenic conditions and during macrophage infection. Surprisingly, intracellular secretion of CpnT was also dependent on both ESX-1 and ESX-4. These secretion defects could be partially rescued by coinfection with wild-type bacteria, indicating that secreted effectors are involved in this process. In summary, our data reveal that three different type VII secretion systems have to be functional in order to observe intracellular secretion of the toxin CpnT.IMPORTANCE For decades, it was believed that the intracellular pathogen M. tuberculosis does not possess toxins. Only fairly recently it was discovered that CpnT is a potent secreted toxin of M. tuberculosis, causing necrotic cell death in host cells. However, until now the secretion pathway remained unknown. In our study, we were able to identify CpnT as a substrate of the mycobacterial type VII secretion system. Pathogenic mycobacteria have up to five different type VII secretion systems, called ESX-1 to ESX-5, which play distinct roles for the pathogen during growth or infection. We were able to elucidate that CpnT is exclusively secreted by the ESX-5 system in bacterial culture. However, to our surprise we discovered that, during infection studies, CpnT secretion relies on intact ESX-1, ESX-4, and ESX-5 systems. We elucidate for the first time the intertwined interplay of three different and independent secretion systems to secrete one substrate during infection.

Thirty-day outcomes for children and adolescents undergoing laparoscopic sleeve gastrectomy at a free-standing children's hospital.

The obesity epidemic continues to affect millions of children and adolescents. Non-surgical options do not result in significant or sustained weight loss; thus bariatric surgery has become increasingly utilized. Limited data exist regarding safety for paediatric bariatric surgery, especially outside of National Institutes of Health (NIH)-funded centres. We hypothesized that the perioperative outcomes of paediatric patients undergoing laparoscopic sleeve gastrectomy (LSG) at our free-standing children's hospital would provide adequate safety profiles. We retrospectively reviewed demographics, comorbidities and 30-d outcomes for all patients who underwent LSG from 2010 to 2015 at a free-standing children's hospital. A total of 105 patients underwent 107 LSG procedures (two revisions). Mean age was 17.2 ± 2.4 years. Male to female ratio was 1:4. The majority were Black (57.1%), followed by White (21.0%) and Hispanic (18.1%). The mean body mass index was 51.0 ± 9.8 kg/m2 . Comorbidities included obstructive sleep apnea (59.0%), hypertension (15.2%), polycystic ovarian disease (16.7% of females only), depression (12.4%) and diabetes (11.4%). Median length of stay was 2.0 d (1-7 d). There were no deaths. Major complications occurred in four patients (3.8%); three required reoperation. Four patients (3.8%) experienced minor complications. Laparoscopic sleeve gastrectomy can be safely performed for children and adolescents at a free-standing children's hospital without NIH-support.

Elevated p21 mRNA level in skeletal muscle of DMD patients and mdx mice indicates either an exhausted satellite cell pool or a higher p21 expression in dystrophin-deficient cells per se.

Abnormalities in proliferation and differentiation of the dystrophin-deficient muscle are a controversial aspect of the pathogenesis of Duchenne muscular dystrophy (DMD). Analyses of molecules involved in cell cycle modulation do not exist in this context. Cells withdrawn from the cell cycle permanently express p21. The fact that p2 1, in contrast to other cell cycle proteins, is not diminished when myotubes are reexposed to growth media, allocates this cyclin-dependent kinase inhibitor a special function. Here we report for the first time statistically increased p21 mRNA levels in dystrophin-deficient muscle tissue. Only 42% of conventional RT-PCRs from six muscle samples of human controls yielded positive results but almost all skeletal muscle biopsy samples (87%) from DMD patients (n=5). For p21 mRNA quantification in murine muscle samples we were able to use the exact real-time TaqMan PCR method due to generally higher p21 mRNA levels than in human muscles. In addition, contamination with fibroblasts can be excluded for the murine samples because they do not demonstrate fibrosis at the age of 350 days but start to lose their regenerative capacity. In accord with the results in humans, we observed p21 mRNA levels in mdx mice that were approx. four times as high as those in control mice. Elevated p21 mRNA level may indicate a shift in cell composition towards differentiated p21 expressing cells as a result of an exhausted pool of undifferentiated, non-p21-expressing satellite cells due to previous cycles of de- and regeneration. Alternatively, dystrophin-deficient cells per se may express higher p21 levels for unknown reasons. Although we cannot distinguish between these possibilities, the eventual transfec tion of a patient's own satellite cells with p21 antisense oligonucleotides may enable the dystrophic process to be influenced.

Relationship of polymorphisms in the renin-angiotensin system and in E-selectin of patients with early severe coronary heart disease.

Previous association studies between angiotensin-converting enzyme (ACE) and angiotensinogen (AGT) polymorphisms and several cardiovascular diseases have reported variable results. Therefore we examined the association of the DNA variants of ACE and AGT with early, severe coronary heart disease (CHD). In addition, we compared the genotypes of both polymorphisms and the recently discovered polymorphism in the E-selectin gene in both patients and an unselected population. This study included 113 patients with severe CHD (50 years old or less) and up to 197 control subjects. The frequencies of the ACE I/D variants were 48% I and 52% D in the controls and 46% I and 54% D in the patients. The frequencies of the AGT-M235T polymorphism were 60.8% M and 39.2% T in controls and 49.1% M and 50.9% T in the patients. The frequencies of the S128R polymorphism of the E-selectin were 91.3% S and 8.7% R in controls and 84.5% S and 15.5% R in the patients. In our studies the DD genotype of ACE was not associated with early severe CHD. We found a correlation between the M235T molecular variant of AGT and the S128R variant of E-selectin to early severe CHD.

Beta1-adrenoceptor gene variations: a role in idiopathic dilated cardiomyopathy?

A substantial body of evidence suggests involvement of the human beta1-adrenoceptor (beta1-AR) gene in the pathophysiology of dilated cardiomyopathy (DCM), a severe heart disease of significant public health impact. Beta1-AR-mediated signal transduction is dramatically altered due to downregulation, resulting in an impairment of myocardial response. The important role of genetic factors in idiopathic dilated cardiomyopathy (IDCM) recently recognized, we analyzed this prime candidate gene for genetic variation in carefully selected patients and controls. In this preliminary study, 18 single nucleotide polymorphisms were observed, 17 of which were located in the N-terminal and C-terminal region of the coding exon, resulting in 7 amino acid exchanges: Ser-49-Gly, Ala-59-Ser, Gly-389-Arg, Arg-399-Cys, His-402-Arg, Thr-404-Ala, and Pro-418-Ala. These mutations resulted in 11 different beta1-AR genotypes. Importantly, the genotypes carrying the Ser-49-Gly mutation in the N-terminus of the molecule in a heterozygous or homozygous form were observed significantly more frequently in the group of IDCM patients. The present results may provide a clue on the molecular mechanisms involved in IDCM, and add moreover interesting information on nature, distribution, and evolutionary aspects of sequence variation in human adrenergic receptor genes.

Opposite effects of high and low frequency rTMS on regional brain activity in depressed patients.

BACKGROUND: High (10-20 Hz) and low frequency (1-5 Hz) repetitive transcranial magnetic stimulation (rTMS) have been explored for possible therapeutic effects in the treatment of neuropsychiatric disorders. As part of a double-blind, placebo-controlled, crossover study evaluating the antidepressant effect of daily rTMS over the left prefrontal cortex, we evaluated changes in absolute regional cerebral blood flow (rCBF) after treatment with 1- and 20-Hz rTMS. Based on preclinical data, we postulated that high frequency rTMS would increase and low frequency rTMS would decrease flow in frontal and related subcortical circuits. METHODS: Ten medication-free, adult patients with major depression (eight unipolar and two bipolar) were serially imaged using (15)O water and positron emission tomography to measure rCBF. Each patient was scanned at baseline and 72 hours after 10 daily treatments with 20-Hz rTMS and 10 daily treatments with 1 Hz rTMS given in a randomized order. TMS was administered over the left prefrontal cortex at 100% of motor threshold (MT). Significant changes in rCBF from pretreatment baseline were determined by paired t test. RESULTS: Twenty-hertz rTMS over the left prefrontal cortex was associated only with increases in rCBF. Significant increases in rCBF across the group of all 10 patients were located in the prefrontal cortex (L > R), the cingulate gyrus (L >> R), and the left amygdala, as well as bilateral insula, basal ganglia, uncus, hippocampus, parahippocampus, thalamus, and cerebellum. In contrast, 1-Hz rTMS was associated only with decreases in rCBF. Significant decreases in flow were noted in small areas of the right prefrontal cortex, left medial temporal cortex, left basal ganglia, and left amygdala. The changes in mood following the two rTMS frequencies were inversely related (r = -.78, p <.005, n = 10) such that individuals who improved with one frequency worsened with the other. CONCLUSIONS: These data indicate that 2 weeks of daily 20-Hz rTMS over the left prefrontal cortex at 100% MT induce persistent increases in rCBF in bilateral frontal, limbic, and paralimbic regions implicated in depression, whereas 1-Hz rTMS produces more circumscribed decreases (including in the left amygdala). These data demonstrate frequency-dependent, opposite effects of high and low frequency rTMS on local and distant regional brain activity that may have important implications for clinical therapeutics in various neuropsychiatric disorders.

A controlled trial of daily left prefrontal cortex TMS for treating depression.

BACKGROUND: Transcranial magnetic stimulation (TMS) is a new technology for noninvasively stimulating the brain. Several studies have suggested that daily stimulation of the left prefrontal cortex with TMS for 2 weeks has probable antidepressant effects. We conducted a parallel-design, double-masked, sham-controlled study to address whether 2 weeks of daily TMS over the left prefrontal cortex has antidepressant activity greater than sham. METHODS: Thirty medication-free adult outpatients with nonpsychotic, major depressive (n = 21) or bipolar (n = 9) (depressed phase) disorder who were in a current major depression (Hamilton Rating Scale for Depression [HRSD] 21-item score of >18) were treated each weekday for 2 weeks. Subjects were randomly assigned to receive either daily active (20 subjects) or sham (10 subjects) stimulation. Additionally, the 20 active subjects were equally divided between slower (5 Hz) and faster (20 Hz) frequency treatment. Antidepressant response was defined as greater than a 50% improvement in the baseline HRSD. RESULTS: Active TMS resulted in significantly more responders (9/20) than did sham (0/10) (chi(2) = 6.42, p <.01). The number of responders did not differ significantly between the two active cells (3/10 faster and 6/10 slower). Expressed as a percent change from baseline, active TMS subjects had significantly greater improvement on the Beck Depression Inventory as well as the Hamilton Anxiety Rating Scale than did those who received sham. CONCLUSIONS: Daily left prefrontal TMS for 2 weeks significantly reduced depression symptoms greater than did sham. The two forms of active TMS treatment did not differ significantly.

Effects of mood and subtype on cerebral glucose metabolism in treatment-resistant bipolar disorder.

BACKGROUND: Functional brain imaging studies in unipolar and secondary depression have generally found decreased prefrontal cortical activity, but in bipolar disorders findings have been more variable. METHODS: Forty-three medication-free, treatment-resistant, predominantly rapid-cycling bipolar disorder patients and 43 age- and gender-matched healthy control subjects had cerebral glucose metabolism assessed using positron emission tomography and fluorine-18-deoxyglucose. RESULTS: Depressed bipolar disorder patients compared to control subjects had decreased global, absolute prefrontal and anterior paralimbic cortical, and increased normalized subcortical (ventral striatum, thalamus, right amygdala) metabolism. Degree of depression correlated negatively with absolute prefrontal and paralimbic cortical, and positively with normalized anterior paralimbic subcortical metabolism. Increased normalized cerebello-posterior cortical metabolism was seen in all patient subgroups compared to control subjects, independent of mood state, disorder subtype, or cycle frequency. CONCLUSIONS: In bipolar depression, we observed a pattern of prefrontal hypometabolism, consistent with observations in primary unipolar and secondary depression, suggesting this is part of a common neural substrate for depression independent of etiology. In contrast, the cerebello-posterior cortical normalized hypermetabolism seen in all bipolar subgroups (including euthymic) suggests a possible congenital or acquired trait abnormality. The degree to which these findings in treatment-resistant, predominantly rapid-cycling patients pertain to community samples remains to be established.

Frequency dependence of antidepressant response to left prefrontal repetitive transcranial magnetic stimulation (rTMS) as a function of baseline cerebral glucose metabolism.

BACKGROUND: Recent studies suggest that both high frequency (10-20 Hz) and low frequency (1 Hz) repetitive transcranial magnetic stimulation (rTMS) have an antidepressant effect in some individuals. Electrophysiologic data indicate that high frequency rTMS enhances neuronal firing efficacy and that low frequency rTMS has the opposite effect. METHODS: We investigated the antidepressant effects of 10 daily left prefrontal 1 Hz versus 20 Hz rTMS with the hypothesis that within a given subject, antidepressant response would differ by frequency and vary as a function of baseline cerebral glucose metabolism. After baseline PET scans utilizing [18F]-Fluorodeoxyglucose, thirteen subjects participated in a randomized crossover trial of 2 weeks of 20 Hz paired with 2 weeks 1 Hz or placebo rTMS. RESULTS: We found a negative correlation between degree of antidepressant response after 1 Hz compared to 20 Hz rTMS (r = -0.797, p < .004). Additionally, better response to 20 Hz was associated with the degree of baseline hypometabolism, whereas response to 1 Hz rTMS tended to be associated with baseline hypermetabolism. CONCLUSIONS: These preliminary results suggest that antidepressant response to rTMS might vary as a function of stimulation frequency and may depend on pretreatment cerebral metabolism. Further studies combining rTMS and functional neuroimaging are needed.

Evidence for quiescent S- and G2-phase cells in human colorectal carcinomas: a flow cytometric study with the Ki-67 antibody.

The expression of certain antigens specific for proliferating cells can be determined simultaneously with cell cycle distribution by means of two-dimensional flow cytometry. In this way, a tumour's growth potential is characterized more precisely than with any one parameter alone. Here we describe such simultaneous measurements of DNA content and labelling with the Ki-67 antibody that distinguishes between cycling and non-cycling cells. Having overcome a number of technical problems we were able to analyse material from 29 biopsies of human colorectal tumours. In a number of cases, Ki-67 negative cells were found with a DNA-content of G0/1 only, whereas all cells with an S- or G2-phase DNA-content were Ki-67 positive. There were other cases in which cells with an S- and G2-phase DNA-content had obviously become quiescent (Ki-67 negative), sometimes even outnumbering the proliferating (Ki- 67 positive) cells in the respective compartments of the cycle. Generally, however, when Ki-67 negative and positive subpopulations were analysed separately it was found that the former had a significantly lower (S + G2)-phase fraction than the latter. There was evidence for a correlation between Ki-67 index and (S + G2)-phase fraction at least in the subgroup of aneuploid tumours. Neither of the two parameters was correlated with stage according to Duke's classification or tumour size. However, a positive correlation was found between the fraction of unlabelled S- and G2-phase cells and tumour size as reflected in the T category.

Assessing the quality of teaching.

Although quality teaching is an integral component of the educational process, it is not easy to define or evaluate. Traditionally, teacher evaluation has focused on student surveys. Peer evaluations are more objective and seem worth the investment. They not only provide a valid and reliable assessment method but have the added advantage of easily being incorporated into faculty development by providing timely, specific feedback. Documentation of teacher accomplishments, both in quantity and quality, can be incorporated into a faculty portfolio and used for promotion, tenure, and salary decisions. Evaluating the quality of courses remains less than satisfactory. Surveying other stakeholders would provide an added perspective that might be valuable to the course. Continued research into curricular outcome measures will be important to assessing quality of courses. Until that time, a combination of student surveys and acquisition of skills will be the most logical methods.

Carrier detection in DMD families with point mutations, using PCR-SSCP and direct sequencing.

Non-isotopic single-strand conformation polymorphism (SSCP) and direct sequencing was used for carrier diagnosis in four families of DMD/BMD patients with previously characterized point mutations, leading to the identification of eight carriers and four non-carriers. When the mutation caused a distinctly altered migration pattern of the single strands, in principle, the SSCP-technique allowed determination of carrier status in the extended family of the probands without direct sequencing. However, because SSCP measures a function of not only the mutation, but of the entire sequence of the PCR product, it can lead to false negative and/or false positive diagnoses due to intronic and exonic sequence heterogeneity in the family. As we discovered this pitfall in one of the reported families, we concluded that for carrier testing the SSCP approach must be performed in essential conjunction with an independent assessment of the mutation site by direct sequencing.

Heterozygous myogenic factor 6 mutation associated with myopathy and severe course of Becker muscular dystrophy.

Myogenic factors (MYF) belong to the basic helix-loop-helix (bHLH) transcription factor family and regulate myogenesis and muscle regeneration. The physiological importance of both functions was demonstrated in homozygous Myf knockout mice and mdx mice. Myf5 and Myod are predominantly expressed in proliferating myoblasts while Myf4 and Myf6 are involved in differentiation of myotubes. In a boy with myopathy and an increase of muscle fibres with central nuclei we detected a heterozygous 387G-->T nucleotide transversion in the MYF6 gene (MIM*159991). Protein-protein interaction of mutant MYF6 was reduced, and DNA-binding potential and transactivation capacity were abolished, thus demonstrating MYF6 haploinsufficiency. The boy's father carried the identical mutation and, in addition, an in-frame deletion of exons 45-47 in his dystrophin gene. This mutation is normally associated with a mild to moderate course of Becker muscular dystrophy but the father suffered from a severe course of Becker muscular dystrophy suggesting MYF6 as a modifier.

A comparison of performance between third-year students completing a pediatric ambulatory rotation on campus vs in the community.

OBJECTIVE: To compare the performance of third-year medical students who completed the ambulatory component of their pediatric rotation in a community setting with the performance of third-year medical students who had their ambulatory experience on campus. METHODS: As part of a pilot project to implement a third-year Multidisciplinary Ambulatory Clerkship, 61 third-year medical students spent 12 weeks rotating through the primary care disciplines of family medicine, internal medicine, and pediatric practitioners' offices at sites distant from the university campus while 127 students remained on campus for their ambulatory experiences in these disciplines. The components of the overall pediatric grade consisted of a clinical performance evaluation in the ambulatory setting (4 weeks), a clinical performance evaluation on a 4-week inpatient rotation, and a grade from a multiple-choice final examination. RESULTS: The overall mean+/-SD final pediatric grade of students receiving their ambulatory pediatrics experience in the Multidisciplinary Ambulatory Clerkship was 86.5+/-3.4 compared with 88.0+/-3.4 for students receiving their ambulatory experience on campus (P<.007). This difference was accounted for by performance on the written final examination. Multidisciplinary Ambulatory Clerkship students had a mean+/-SD score of 78.9+/-8.3 and a failure rate of 18% compared with a mean score of 83.7+/-8.1 and failure rate of 3.9% for students who remained on campus for their ambulatory experience (P<.001 for both comparisons). No differences were noted between the 2 groups on their clinical performance evaluations for their ambulatory or inpatient experiences. CONCLUSIONS: These data suggest a difference in the learning experience between students receiving their pediatric ambulatory experience in the community vs on campus. Differences in exposure to structured learning experiences that occurred more frequently on campus might account for some of the difference in final examination results. Development of a standardized, structured learning experience across community sites would seem to be an appropriate means of enhancing learning in the community setting.

The relationship between preceptor expectations and student performance on 2 pediatric objective structured clinical examination stations.

BACKGROUND: We designed 2 pediatric objective structured clinical examination stations, 1 anemia case associated with lead exposure and 1 failure-to-gain-weight case associated with extended breast-feeding, to evaluate third-year medical students who had studied in pediatric community preceptors' offices as part of a 12-week multidisciplinary ambulatory clerkship rotation. OBJECTIVE: To examine the relationship between preceptor expectations and student performance on these 2 objective structured clinical examination stations. METHODS: To elicit community preceptors' expectations of student performance, we constructed a 46-item survey replicating checklists filled out by simulated patients evaluating student performance on the objective structured clinical examination stations. The percentage agreement among preceptors for each checklist item as well as the percentage agreement between preceptor responses and student responses on each checklist item were calculated. A summary score of preceptor responses across all checklist items and a summary score for student responses across all checklist items on each station were calculated. The correlation coefficients between preceptor and student summary scores were then examined. RESULTS: Fifty-nine preceptor surveys were mailed and 38 were returned (64% response rate). Data were usable from 37 surveys. Eighty-nine percent (33 of 37)of the preceptors agreed that a third-year clerkship student should have the knowledge to care for the patient with anemia and 92% (34 of 37)of the preceptors agreed similarly for the growth-delay case. Agreement among preceptors on individual checklist items varied widely for both cases. Fifty-seven students studied at the anemia station and 34 students studied at the growth-delay station. The mean+/-SD agreement across the 26 items on the anemia case between preceptor responses and student responses was 62%+/-23% and, for the 21 items on the growth-delay case, 60%+/-17%. The mean+/-SD preceptor summary score for the anemia case was 17.4+/-3.8 (maximum, 26) and 16.0+/-3.6 (maximum, 21) for the growth-delay case. The mean student score on the anemia case was 15.5+/-3.7 (maximum, 26) and, for the growth-delay case, 10.0+/-4.5 (maximum, 21). The Pearson correlation coefficient between the preceptor and student scores on the anemia case was 0.19 (P=.15), and for the growth-delay case,-0.41 (P=.06). CONCLUSIONS: These data suggest community preceptors agree on topic areas in which students should be clinically competent. There was, however, considerable variation in agreement among preceptors about what preceptors believe students should be able to do and how the students actually perform. The overall percentage agreement between preceptor expectations and student performance appears to be no better than chance.

Genomic organization of the human excitatory amino acid transporter gene GLT-1.

We present the genomic structure of the human glutamate transporter GLT-1 coding region, the intronic sequences adjacent to the exons, and oligonucleotide primer sequences for single strand conformational analysis. The exon-intron boundaries were determined using long-distance PCR and direct sequencing. The human GLT-1 coding region is composed of 10 exons spanning > 50 kb of genomic DNA. The exons range from 127 to 251 bp in length. The intron lengths vary considerably from 2.2 kb to > 15 kb. These data provide the basis for implementing a comprehensive screen for genetic alterations in the human GLT-1 gene using genomic DNA as a template.

Mapping transcranial magnetic stimulation (TMS) fields in vivo with MRI.

Transcranial magnetic stimulation (TMS) is a non-invasive technique for investigating brain function that uses pulsed magnetic fields created by special coils to induce localized neuronal depolarization. Despite the technique's expanding application, the exact magnetic field produced by TMS coils have never been directly measured in human subjects. Using a standard 1.5T MR scanner and TMS coils constructed from non magnetic materials, we have obtained 3D maps of the magnetic field created by TMS coils in human volunteers. Further, we mapped the combined field of two coils and demonstrated that combinations of coils might be used to focus the magnetic field to achieve improved stimulation patterns and, perhaps, reach areas out of reach of single coils.