Profiling learning strategies of medical students: A person-centered approach.

BACKGROUND: Students within a cohort might employ unique subsets of learning strategies (LS) to study. However, little research has aimed to elucidate subgroup-specific LS usage among medical students. Recent methodological developments, particularly person-centred approaches such as latent profile analysis (LPA), offer ways to identify relevant subgroups with dissimilar patterns of LS use. In this paper, we apply LPA to explore subgroups of medical students during preclinical training in anatomy and examine how these patterns are linked with learning outcomes. METHODS: We analysed the LS used by 689 undergraduate, 1st and 2nd-year medical students across 6 German universities who completed the short version of the Learning Strategies of University Students (LIST-K) questionnaire, and answered questions towards external criteria such as learning resources and performance. We used the thirteen different LS facets of the LIST-K (four cognitive, three metacognitive, three management of internal and three management of external resources) as LPA indicators. RESULTS: Based on LPA, students can be grouped into four distinct learning profiles: Active learners (45% of the cohort), collaborative learners (17%), structured learners (29%) and passive learners (9%). Students in each of those latent profiles combine the 13 LS facets in a unique way to study anatomy. The profiles differ in both, the overall level of LS usage, and unique combinations of LS used for learning. Importantly, we find that the facets of LS show heterogeneous and subgroup-specific correlations with relevant outcome criteria, which partly overlap but mostly diverge from effects observed on the population level. CONCLUSIONS: The effects observed by LPA expand results from variable-centered efforts and challenge the notion that LS operate on a linear continuum. These results highlight the heterogeneity between subgroups of learners and help generate a more nuanced interpretation of learning behaviour. Lastly, our analysis offers practical implications for educators seeking to tailor learning experiences to meet individual student needs.

Evaluation of the (clinical) relevance of gross anatomical teaching for dental students and practicing oral surgeons in Berlin.

Knowledge of human anatomy is an important scientific basis for every dentist, and the adequacy of its provision by anatomy faculties is therefore constantly being adapted. Students' motivation to comprehend knowledge is a fundamental prerequisite and has been shown to increase when the relevance of the subject is clear. This study examines dental students' perceptions of the relevance of the anatomy curriculum, with particular reference to clinical practice and the dissection course, in conjunction with the perceptions of surgically specialized dentists. The distinctions between the participating groups concerning their perceived relevance are being discussed particularly regarding their applicability to improve the anatomical education of dental students. The overall aim is to find and highlight aspects of the clinical use of anatomical knowledge towards students, which they are unable to apprehend yet. A questionnaire consisting of information on demographics and professional competence as well as a Likert-style section was administered to dental students at Charité Universitätsmedizin in 2019/2020 (n = 322, 84.96%) and a modified version with congruent questions was administered retrospectively to oral surgeons practicing in Berlin in 2020 (n = 81, 63.3%). A Wilcoxon rank-sum test was used to examine differences in responses between the cohorts. Demographic data and professional competence were correlated with the participants' ratings of the given statements using Spearman's rank correlation coefficient. Both groups of respondents expressed a high level of appreciation of the overall relevance of anatomical knowledge, valued the teaching of all human anatomy for dental education, and recognized the relevance of anatomical knowledge for clinical practice. The importance of the dissection course was also rated relatively high by both groups of respondents. The overall appreciation of anatomical knowledge was more prevalent among oral surgeons. There were few correlations between demographics in our findings, showing slightly higher motivation in female students and a growing appreciation of anatomical knowledge as students qualification progressed. The demographics and post-qualification experience of the participating oral surgeons did not influence their perceptions. The results of this survey suggest that there is potential to increase the motivation of dental students to learn anatomy by emphasizing clinical relevance, as perceived by experienced practitioners, during the anatomy curriculum.

Comparison of the study performance of dental students in different subfields of neuroanatomy at the Charité - Universitätsmedizin Berlin and evaluation of the neuroanatomy course in the fourth preclinical semester.

INTRODUCTION: The dentist's main working area is the head and neck region, which is innervated by the cranial nerves. On a daily basis, dentists must administer local anaesthesia to ensure pain-free treatment and differentiate between dental pain and neuropathies to avoid mistreatment. Therefore, neuroanatomical training, especially on the cranial nerves, is of immense importance for clinical practice. In order to adopt the curriculum, it is essential to constantly evaluate the quality of the training and to investigate whether there is a correlation between the students' performance and the relevance of the subfields to their work. MATERIAL AND METHODS: To address this issue, the results of MC exams in the neuroanatomy course for dental students at Charité-Universitätsmedizin Berlin from winter semester 2014/2015 to winter semester 2019/2020 were analysed. Each question was assigned to a specific subfield of neuroanatomy. We then compared cranial nerves and cranial nerve nuclei (clinically relevant) with the remaining subfields (clinically less/not relevant) to investigate whether students performed better in anatomy subfields that are more aligned with the clinical practice of a dentist. We also conducted an anonymous survey (n=201) of the dental students. RESULTS: From winter semester 2014/2015 to winter semester 2019/2020, students performed significantly (***, p< 0.001) better on the clinically relevant questions of the MC examination than on the less/not clinically relevant questions. However, when looking at each of the eleven semesters separately, only three semesters actually performed significantly better on the clinically relevant questions. Our survey also showed that students perceived the subfield of cranial nerves and cranial nerve nuclei to be the most relevant and studied it more intensively out of their own interest. DISCUSSION: The study showed that students perceived the subfield of cranial nerves and cranial nerve nuclei to be the most relevant. However, there was no direct correlation between student performance and clinically relevant questions. Using student performance alone as an indicator of relevance is not optimal, as factors such as motivation to learn can have a significant impact. CONCLUSION: Greater clinical relevance influences what students learn more intensively out of their own interest, but does not influence the results of the MC examination in favour of the subspecialty. Based on the available evidence, it is recommended that the structure of the neuroanatomy course be reconsidered.

Template-based target point localization for occlusion of the middle meningeal artery during evacuation of space-occupying chronic subdural hematoma.

OBJECTIVE: Endovascular middle meningeal artery (MMA) occlusion may help reduce the risk of recurrence after burr hole evacuation of chronic subdural hematoma (cSDH) but carries an additional periprocedural risk and remains hampered by logistical and financial requirements. In this study, the authors aimed to describe a simple and fast technique for preoperative MMA localization to permit burr hole cSDH evacuation and MMA occlusion through the same burr hole. METHODS: The authors performed a preclinical anatomical and prospective clinical study, followed by a retrospective feasibility analysis. An anatomical cadaver study with 33 adult human skulls (66 hemispheres) was used to localize a suitable frontal target point above the pterion, where the MMA can be accessed via burr hole trephination. Based on anatomical landmark measurements, the authors designed a template for projected localization of this target point onto the skin. Next, the validity of the template was tested using image guidance in 10 consecutive patients undergoing elective pterional craniotomy, and the feasibility of the target point localization for cSDH accessibility was determined based on hematoma localization in 237 patients who were treated for a space-occupying cSDH in the authors' department between 2014 and 2018. RESULTS: In the anatomical study, the mean perpendicular distance from the zygomatic process to the target point in the frontoparietal bone was 4.1 cm (95% CI 4-4.2 cm). The mean length along the upper margin of the zygomatic process from the middle of the external auditory canal to the point of the perpendicular distance was 2.3 cm (95% CI 2.2-2.4 cm). The template designed according to these measurements yielded high agreement between the template-based target point and the proximal MMA groove inside the frontoparietal bone (right 90.9%; left 93.6%). In the clinical validation, we noted a mean distance of 4 mm (95% CI 2.1-5.9 mm) from the template-based target point to the actual MMA localization. The feasibility analysis yielded that 95% of all cSDHs in this cohort would have been accessible by the new frontal burr hole localization. CONCLUSIONS: A template-based target point approach for MMA localization may serve as a simple, fast, reliable, and cost-effective technique for surgical evacuation of space-occupying cSDHs with MMA obliteration through the same burr hole in a single setting.

Evaluation of a 3D-MC examination format in anatomy.

INTRODUCTION: The quality of education in medical anatomy is a fundamental pillar of good clinical practice. Current reforms of the medical curriculum have resulted in major methodological changes in the teaching and testing of anatomy. A number of recent studies have however described a decrease in positive metrics of anatomical knowledge among students so taught. It has been suggested that the reduced anatomical knowledge measured in these studies may endanger patient safety. As proxy measures of exam quality, evaluation of the levels of students 'achievement in the examinations, assessment of the subjectively perceived level of question difficulty and analysis of exam satisfaction are each suitable parameters of investigation of medical education. MATERIAL AND METHODS: To address these issues with regard to medical education at the Charité-Universitätsmedizin Berlin, we have analyzed students' levels of achievement in the anatomical Three Dimensional Multiple Choice (hereafter, 3D-MC)-examination of 2,015 students matriculated in medical studies from Summer Semester of 2014 through Summer Semester of 2017. We either compared students' achievement levels of identical 3D-MC questions using models or prepared anatomical specimen. Furthermore, we have analyzed the type and frequency of cognitive levels used in the anatomical questions in relation to the students' level of achievement. Finally, we conducted an anonymous survey to measure students' (n = 207) and instructors' (n = 16) satisfaction with the 3D-MC-examination in comparison to other employed anatomical testing strategies. RESULTS: Students' achievement is significantly enhanced with anatomical questions using models relative to those utilizing anatomical specimen. Over 80% of the anatomical questions in the 3D-MC-examinations assessed the lowest cognitive levels and higher cognitive question levels were accompanied by a significant decrease of the levels of students' performance. Our survey further revealed that both, students and instructors preferred the practical examinations in anatomy and that the difficulty levels of the 3D-MC-examination was perceived as being the lowest in comparison to the other anatomical testing strategies. DISCUSSION: Testing levels of anatomical understanding using anatomical models is not comparable to human specimen, and thus using specimen before models should be preferred to learn and test close to an authentic medical situation. The application of anatomical models and low cognitive question levels in the examination reduces the subjectively perceived level of difficulty, encourages superficial learning, and therefore decreases the retention of anatomical knowledge. CONCLUSION: Although students and instructors prefer practical examinations in anatomy, the current development does not reflect these results. Therefore, it would be recommendable to rethink the development of anatomical testing strategies based on the existing evidence.

Challenges and implementation of the German maternity protection act for female medical students in macroscopic anatomical education.

Background: Recent decades have seen controversial discussions on the validity of dissection courses in medical education, with alternative programs tested for various reasons. On April 1, 2015 the classification of formaldehyde as a hazardous substance was upgraded by the EU, leding to some universities precluding the participation of pregnant and breastfeeding students in dissection course. However, the revision to the Maternity Protection Act, implemented in Germany on January 1, 2018, now protects student mothers from being disadvantaged in their studies as a consequence of their pregnancy or breastfeeding. Therefore, universities must offer alternatives to dissection courses using formaldehyde to these female students. Project description: As an alternative to regular dissection courses, which use the abovementioned chemical, the Centre for Anatomy at Charité has opted for developing dedicated courses for student mothers. These new courses use plastinated prosection material instead of formalin-treated cadavers of body donors. As the core of the anatomical education takes place during the third and fourth semester in the current curriculum of human medicine at Charité the alternative courses are limited to those two semesters. Additionally, alternative exams at the end of both semesters had to be developed. The alternative courses were designed to offer pregnant and breastfeeding students a study program as close as possible to the one in which their peers learn human anatomy. Results: For the new courses, plastinates had to be produced and further specimens are still needed. Additionally required sets of bones, models and radiological images were readily available at the Centre for Anatomy. The planning and conceptualization of the courses took half a year of intense preparation. The courses for the third and fourth semester were first running during summer semester 2017. There is a clear demand for courses among pregnant and breastfeeding students. At least 5 student participants per course were registered, corresponding to every fortieth female student in their semester cohorts. The highest number of student participants was 13 in one course so far. The performances of the participants in the anatomical examinations were matching that of students attending the regular courses. Discussion: The alternative macroscopic anatomy courses enable the implementation of the revised Maternity Protection Act. The targeted student group is highly satisfied with the offered alternative courses. Considering the number of participants and their examination performance so far, the Centre for Anatomy regards the efforts involved in planning and implementing the courses as justified. The courses allow pregnant and breastfeeding students to address the same anatomical themes at the same time as their fellow students. However, due to restricted flexibility of plastinates and because students cannot prepare specific anatomical structures independently the scope of topographic learning is limited. That being said, well-produced plastinates can display anatomical structures which often cannot be dissected in regular courses. The alternative macroscopic anatomy courses using plastinates constitute suitable alternatives to the regular dissection courses with formalin-treated cadavers for pregnant and breastfeeding students.

An Alternative Exon of CAPS2 Influences Catecholamine Loading into LDCVs of Chromaffin Cells.

The calcium-dependent activator proteins for secretion (CAPS) are priming factors for synaptic and large dense-core vesicles (LDCVs), promoting their entry into and stabilizing the release-ready state. A modulatory role of CAPS in catecholamine loading of vesicles has been suggested. Although an influence of CAPS on monoamine transporter function and on vesicle acidification has been reported, a role of CAPS in vesicle loading is disputed. Using expression of naturally occurring splice variants of CAPS2 into chromaffin cells from CAPS1/CAPS2 double-deficient mice of both sexes, we show that an alternative exon of 40 aa is responsible for enhanced catecholamine loading of LDCVs in mouse chromaffin cells. The presence of this exon leads to increased activity of both vesicular monoamine transporters. Deletion of CAPS does not alter acidification of vesicles. Our results establish a splice-variant-dependent modulatory effect of CAPS on catecholamine content in LDCVs.SIGNIFICANCE STATEMENT The calcium activator protein for secretion (CAPS) promotes and stabilizes the entry of catecholamine-containing vesicles of the adrenal gland into a release-ready state. Expression of an alternatively spliced exon in CAPS leads to enhanced catecholamine content in chromaffin granules. This exon codes for 40 aa with a high proline content, consistent with an unstructured loop present in the portion of the molecule generally thought to be involved in vesicle priming. CAPS variants containing this exon promote serotonin uptake into Chinese hamster ovary cells expressing either vesicular monoamine transporter. Epigenetic tuning of CAPS variants may allow modulation of endocrine adrenaline and noradrenaline release. This mechanism may extend to monoamine release in central neurons or in the enteric nervous system.

Balance of Go1α and Go2α expression regulates motor function via the striatal dopaminergic system.

The heterotrimeric G-protein Go with its splice variants, Go1α and Go2α, seems to be involved in the regulation of motor function but isoform-specific effects are still unclear. We found that Go1α-/- knockouts performed worse on the rota-rod than Go2α-/- and wild-type (WT) mice. In Go1+2α-/- mice motor function was partially recovered. Furthermore, Go1+2α-/- mice showed an increased spontaneous motor activity. Compared to wild types or Go2α-/- mice, Go1+2α-/- mice developed increased behavioural sensitization following repetitive cocaine treatment, but failed to develop conditioned place preference. Analysis of dopamine concentration and expression of D1 and D2 receptors unravelled splice-variant-specific imbalances in the striatal dopaminergic system: In Go1α-/- mice dopamine concentration and vesicular monoamine uptake were increased compared to wild types. The expression of the D2 receptor was higher in Go1α-/- compared to wild type littermates, but unchanged in Go2α-/- mice. Deletion of both Go1α and Go2α re-established both dopamine and D2 receptor levels comparable to those in the wild-type. Cocaine treatment had no effect on the ratio of D1 receptor to D2 receptor in Go1+2α-/- mutants, but decreased this ratio in Go2α-/- mice. Finally, we observed that the deletion of Go1α led to a threefold higher striatal expression of Go2α. Taken together our data suggest that a balance in the expression of Go1α and Go2α sustains normal motor function. Deletion of either splice variant results in divergent behavioural and molecular alterations in the striatal dopaminergic system. Deletion of both splice variants partially restores the behavioural and molecular changes. Open Data: Materials are available on https://cos.io/our-services/open-science-badges/ https://osf.io/93n6m/.

Do they know too little? An inter-institutional study on the anatomical knowledge of upper-year medical students based on multiple choice questions of a progress test.

The depth of medical students' knowledge of human anatomy is often controversially discussed. In particular, members of surgical disciplines raise concerns regarding deficits in the factual anatomical and topographical knowledge of upper-year students. The question often raised is whether or not medical students have sufficient anatomical and topographical knowledge when they graduate from medical school. Indeed, this question is highly relevant for curricular planners. Therefore, we have addressed it by evaluating the performance of students in the 5th and 6th years of their studies on anatomical multiple choice questions from the Berlin Progress Test Medicine performed at 10 German university medical schools. Results were compared to a reference based on a standard setting (modified Angoff-procedure). The reference was established independently by 5 panels of anatomists at different universities across Germany. As the ratings were independent of university affiliation, teaching-experience or training of the anatomists, an overall cut off score could be calculated which corresponded to 60.4% correct answers for the question set used in this study. In the progress test, on average only 29.9% of the students' answers were correct, reflecting that the performance was significantly below the expected standard. On the basis of the test results it remained unclear whether acquisition or retention of anatomical information was insufficient. Further evaluation by item characteristics revealed that the students had major difficulty in applying their theoretical knowledge to practical problems in the context of a clinical setting. Thus, our results reveal deficits in the anatomical knowledge of medical students in their final years. Therefore medical curricula should not only focus on enhancing the acquisition and retention of core anatomical knowledge, but aim at improving skills applying this in a clinical setting.

The α-subunit of the trimeric GTPase Go2 regulates axonal growth.

The Goα splice variants Go1α and Go2α are subunits of the most abundant G-proteins in brain, Go1 and Go2. Only a few interacting partners binding to Go1α have been described so far and splice variant-specific differences are not known. Using a yeast two-hybrid screen with constitutively active Go2α as bait, we identified Rap1GTPase activating protein (Rap1GAP) and Girdin as interacting partners of Go2α, which was confirmed by co-immunoprecipitation. Comparison of subcellular fractions from brains of wild type and Go2α-/- mice revealed no differences in the overall expression level of Girdin or Rap1GAP. However, we found higher amounts of active Rap1-GTP in brains of Go2α deficient mutants, indicating that Go2α may increase Rap1GAP activity, thereby effecting the Rap1 activation/deactivation cycle. Rap1 has been shown to be involved in neurite outgrowth and given a Rap1GAP-Go2α interaction, we found that the loss of Go2α affected axonal outgrowth. Axons of cultured cortical and hippocampal neurons prepared from embryonic Go2α-/- mice grew longer and developed more branches than those from wild-type mice. Taken together, we provide evidence that Go2α regulates axonal outgrowth and branching.

A speedy recovery: amphetamines and other therapeutics that might impact the recovery from brain injury.

PURPOSE OF REVIEW: Traumatic brain injury is the leading cause of death in young patients and stroke is the leading cause of major disability in elderly patients. Both injuries are often followed by cerebral plasticity changes and increased intracerebral pressure (ICP). Aim of the review is to present current knowledge about amphetamines and other therapeutics concerning the recovery of brain injury based on contemporary findings. RECENT FINDINGS: On the one hand beneficial effects for cognitive and physical outcome after brain injury by coupling amphetamine with physical therapy could be demonstrated; on the other hand its efficacy was shown in only two out of 10 clinical trials. SUMMARY: Impairment after brain injury is reduced if adequate early treatment is established. On the basis of current findings amphetamine after brain injury cannot be recommended. In patients with an increased ICP the maintenance of an adequate cerebral perfusion pressure is required. In patients with increased ICP under controlled ventilation, the combination of ketamine with, for example, a short-acting benzodiazepine and opioid or methohexital is equally well tolerated. Ketamine decreases ICP without lowering blood pressure and cerebral perfusion pressure. Its neuroprotective property might reduce the exacerbation of brain injury following N-methyl-D-aspartate-receptor activation, neuronal apoptosis and systemic inflammatory responses.

Neurotoxic phospholipases directly affect synaptic vesicle function.

Snake neurotoxic phospholipases (SPAN) exclusively affect pre-synaptic nerve terminals where they lead to a block of neurotransmission by not fully understood mechanisms. Here, we report that the SPANs, taipoxin and paradoxin, in nanomolar concentrations directly dissociate the synaptophysin/synaptobrevin (Syp/Syb) complex on isolated synaptic vesicles in the presence of synaptosomal cytosol. The phospholipase activity of SPANs depends on Ca(2+) but the dissociation of the Syp/Syb complex does not require Ca(2+). Ca(2+) (100 µM free) alone also dissociates the Syp/Syb complex in the presence of cytosol. Treatment with SPANs disturbs the lipid raft association of synaptophysin and synaptobrevin comparable to cholesterol depletion by ß-methyl-cyclodextrin while Ca(2+) alone has no effect. SPANs but not Ca(2+) directly inhibit vesicular uptake of serotonin and glutamate. It is concluded that SPANs directly affect vesicular properties independent from their Ca(2+) -dependent phospholipase activity. SPANs and Ca(2+) dissociate the Syp/Syb complex as a prerequisite for exocytosis. SPANs also prevent the filling of synaptic vesicles thereby adding to the inhibition of neurotransmission.

Amphetamine regulates NR2B expression in Go2α knockout mice and thereby sustains behavioral sensitization.

The α-subunit of Go2 is a regulator of dopamine (DA) homeostasis. Deletion of the protein results in an imbalance of the direct and indirect DA pathway by reducing D1 and increasing D2 receptors. As a result, cocaine-induced behavioral sensitization is abolished. Here we show that repeated amphetamine injections in Go2α-/- mice induced a similar D1/D2 receptor ratio shift as cocaine but surprisingly the knockouts developed normal behavioral sensitization. DA receptor signaling following either cocaine or amphetamine treatment was also similar in Go2α-/- mice suggesting another mechanism involved in the differential behavioral response. Evidence is increasing that DA-glutamate interactions in the striatum determine psychostimulant action. In this line, repeated amphetamine injections led to a twofold increase in the amount of the NMDA receptor subunit NR2B in Go2α-/- mice resulting in an enhanced inhibition of the indirect DA pathway. This effect is not seen after cocaine treatment. Furthermore, amphetamine but not cocaine treatment maintained the ratio between the glutamate receptor mGluR1/5 interacting proteins Homer and Homer1a in the knockouts thereby sustaining the direct pathway. We conclude that amphetamine provokes behavioral sensitization in Go2α-/- mice by an enhanced inhibition of the indirect pathway without disturbing the direct pathway thereby overcoming the imbalance in the DArgic system.

Synaptic and vesicular coexistence of VGLUT and VGAT in selected excitatory and inhibitory synapses.

The segregation between vesicular glutamate and GABA storage and release forms the molecular foundation between excitatory and inhibitory neurons and guarantees the precise function of neuronal networks. Using immunoisolation of synaptic vesicles, we now show that VGLUT2 and VGAT, and also VGLUT1 and VGLUT2, coexist in a sizeable pool of vesicles. VGAT immunoisolates transport glutamate in addition to GABA. Furthermore, VGLUT activity enhances uptake of GABA and monoamines. Postembedding immunogold double labeling revealed that VGLUT1, VGLUT2, and VGAT coexist in mossy fiber terminals of the hippocampal CA3 area. Similarly, cerebellar mossy fiber terminals harbor VGLUT1, VGLUT2, and VGAT, while parallel and climbing fiber terminals exclusively contain VGLUT1 or VGLUT2, respectively. VGLUT2 was also observed in cerebellar GABAergic basket cells terminals. We conclude that the synaptic coexistence of vesicular glutamate and GABA transporters allows for corelease of both glutamate and GABA from selected nerve terminals, which may prevent systemic overexcitability by downregulating synaptic activity. Furthermore, our data suggest that VGLUT enhances transmitter storage in nonglutamatergic neurons. Thus, synaptic and vesicular coexistence of VGLUT and VGAT is more widespread than previously anticipated, putatively influencing fine-tuning and control of synaptic plasticity.

Quantitative comparison of glutamatergic and GABAergic synaptic vesicles unveils selectivity for few proteins including MAL2, a novel synaptic vesicle protein.

Synaptic vesicles (SVs) store neurotransmitters and release them by exocytosis. The vesicular neurotransmitter transporters discriminate which transmitter will be sequestered and stored by the vesicles. However, it is unclear whether the neurotransmitter phenotype of SVs is solely defined by the transporters or whether it is associated with additional proteins. Here we have compared the protein composition of SVs enriched in vesicular glutamate (VGLUT-1) and GABA transporters (VGAT), respectively, using quantitative proteomics. Of >450 quantified proteins, approximately 50 were differentially distributed between the populations, with only few of them being specific for SVs. Of these, the most striking differences were observed for the zinc transporter ZnT3 and the vesicle proteins SV2B and SV31 that are associated preferentially with VGLUT-1 vesicles, and for SV2C that is associated mainly with VGAT vesicles. Several additional proteins displayed a preference for VGLUT-1 vesicles including, surprisingly, synaptophysin, synaptotagmins, and syntaxin 1a. Moreover, MAL2, a membrane protein of unknown function distantly related to synaptophysins and SCAMPs, cofractionated with VGLUT-1 vesicles. Both subcellular fractionation and immunolocalization at the light and electron microscopic level revealed that MAL2 is a bona-fide membrane constituent of SVs that is preferentially associated with VGLUT-1-containing nerve terminals. We conclude that SVs specific for different neurotransmitters share the majority of their protein constituents, with only few vesicle proteins showing preferences that, however, are nonexclusive, thus confirming that the vesicular transporters are the only components essential for defining the neurotransmitter phenotype of a SV.

Age-dependent axonal expression of potassium channel proteins during development in mouse hippocampus.

The development of the hippocampal network requires neuronal activity, which is shaped by the differential expression and sorting of a variety of potassium channels. Parallel to their maturation, hippocampal neurons undergo a distinct development of their ion channel profile. The age-dependent dimension of ion channel occurrence is of utmost importance as it is interdependently linked to network formation. However, data regarding the exact temporal expression of potassium channels during postnatal hippocampal development are scarce. We therefore studied the expression of several voltage-gated potassium channel proteins during hippocampal development in vivo and in primary cultures, focusing on channels that were sorted to the axonal compartment. The Kv1.1, Kv1.2, Kv1.4, and Kv3.4 proteins showed a considerable temporal variation of axonal localization among neuronal subpopulations. It is possible, therefore, that hippocampal neurons possess cell type-specific mechanisms for channel compartmentalization. Thus, age-dependent axonal sorting of the potassium channel proteins offers a new approach to functionally distinguish classes of hippocampal neurons and may extend our understanding of hippocampal circuitry and memory processing.

Synaptic PRG-1 modulates excitatory transmission via lipid phosphate-mediated signaling.

Plasticity related gene-1 (PRG-1) is a brain-specific membrane protein related to lipid phosphate phosphatases, which acts in the hippocampus specifically at the excitatory synapse terminating on glutamatergic neurons. Deletion of prg-1 in mice leads to epileptic seizures and augmentation of EPSCs, but not IPSCs. In utero electroporation of PRG-1 into deficient animals revealed that PRG-1 modulates excitation at the synaptic junction. Mutation of the extracellular domain of PRG-1 crucial for its interaction with lysophosphatidic acid (LPA) abolished the ability to prevent hyperexcitability. As LPA application in vitro induced hyperexcitability in wild-type but not in LPA(2) receptor-deficient animals, and uptake of phospholipids is reduced in PRG-1-deficient neurons, we assessed PRG-1/LPA(2) receptor-deficient animals, and found that the pathophysiology observed in the PRG-1-deficient mice was fully reverted. Thus, we propose PRG-1 as an important player in the modulatory control of hippocampal excitability dependent on presynaptic LPA(2) receptor signaling.

Ca2+-dependent activator proteins of secretion promote vesicular monoamine uptake.

Ca(2+)-dependent activator proteins of secretion (CAPS) 1 and 2 are essential regulators of synaptic vesicle and large dense core vesicle priming in mammalian neurons and neuroendocrine cells. CAPS1 appears to have an additional and as yet unexplained function in vesicular catecholamine uptake or storage as CAPS1-deficient chromaffin cells exhibit strongly reduced vesicular catecholamine levels. Here we describe a role of CAPS proteins in vesicular monoamine uptake. Both CAPS1 and CAPS2 promote monoamine uptake and storage mediated by the vesicular monoamine transporters VMAT1 and VMAT2. Monoamine uptake of vesicular preparations from embryonic brains of CAPS1 deletion mutants is decreased as compared with corresponding preparations from wild type littermates, and anti-CAPS1 or anti-CAPS2 antibodies inhibit monoamine sequestration by synaptic vesicles from adult mouse brain. In addition, overexpression of CAPS1 or CAPS2 enhances vesicular monoamine uptake in Chinese hamster ovary cells that stably express VMAT1 or VMAT2. CAPS function has been linked to the heterotrimeric GTPase G(o), which modulates vesicular monoamine uptake. We found that the expression of CAPS1 is decreased in brain membrane preparations from mice lacking G(o2)alpha, which may explain the reduced monoamine uptake by G(o2)alpha-deficient synaptic vesicles. Accordingly, anti-CAPS1 antibodies do not further reduce monoamine uptake by G(o2)alpha-deficient synaptic vesicles, whereas antibodies directed against CAPS2, whose expression is not altered in G(o2)alpha-deficient brain, still reduce monoamine uptake into G(o2)alpha-deficient vesicles. We conclude that CAPS proteins are involved in optimizing vesicular monoamine uptake and storage mediated by VMAT1 and VMAT2.

Deletion of Go2alpha abolishes cocaine-induced behavioral sensitization by disturbing the striatal dopamine system.

The alpha-subunits of the trimeric Go class of GTPases, comprising the splice variants Go1alpha and Go2alpha, are abundantly expressed in brain and reside on both plasma membrane and synaptic vesicles. Go2alpha is involved in the vesicular storage of monoamines but its physiological relevance is still obscure. We now show that genetic depletion of Go2alpha reduces motor activity induced by dopamine-enhancing drugs like cocaine, as repeated injections of cocaine fail to provoke behavioral sensitization in Go2alpha(-/-) mice. In Go2alpha(-/-) mice, D1 receptor signaling in the striatum is attenuated due to a reduced expression of Golf alpha and Gs alpha. Following cocaine treatment, Go2alpha(-/-) mice have lower D1 and higher D2 receptor amounts compared to wild-type mice. The lack of behavioral sensitization correlates with reduced dopamine levels in the striatum and decreased expression of tyrosine hydroxylase. One reason for the neurochemical changes may be a reduced uptake of monoamines by synaptic vesicles from Go2alpha(-/-) mice as a consequence of a lowered set point for filling. We conclude that Go2alpha optimizes vesicular filling which is instrumental for normal dopamine functioning and for the development of drug-induced behavioral sensitization.

Regulation of monoamine oxidase A by circadian-clock components implies clock influence on mood.

The circadian clock has been implicated in addiction and several forms of depression [1, 2], indicating interactions between the circadian and the reward systems in the brain [3-5]. Rewards such as food, sex, and drugs influence this system in part by modulating dopamine neurotransmission in the mesolimbic dopamine reward circuit, including the ventral tegmental area (VTA) and the ventral striatum (NAc). Hence, changes in dopamine levels in these brain areas are proposed to influence mood in humans and mice [6-10]. To establish a molecular link between the circadian-clock mechanism and dopamine metabolism, we analyzed the murine promoters of genes encoding key enzymes important in dopamine metabolism. We find that transcription of the monoamine oxidase A (Maoa) promoter is regulated by the clock components BMAL1, NPAS2, and PER2. A mutation in the clock gene Per2 in mice leads to reduced expression and activity of MAOA in the mesolimbic dopaminergic system. Furthermore, we observe increased levels of dopamine and altered neuronal activity in the striatum, and these results probably lead to behavioral alterations observed in Per2 mutant mice in despair-based tests. These findings suggest a role of circadian-clock components in dopamine metabolism highlighting a role of the clock in regulating mood-related behaviors.