Science topic

Neurobiology - Science topic

The study of the structure, growth, activities, and functions of NEURONS and the NERVOUS SYSTEM.
Questions related to Neurobiology
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I will really appreciate it if somebody could share the software. My lab owns the stimulator but when we acquired it we didn't get the software nor the drivers. Grass doesn't exist anymore and "Natus" which is the company that bought it told me that "S88X" is a discontinued product and they do not longer provide support for it.
Its crucial for us to control the device from a PC.
Thanks in advance
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Hi David Martínez-Vargas Janelle M Fine Eric Kenji Lee ! Do any of you still have the Grass S88X Stimulator driver and software? If yes, can you please send it to me at apoorvar@umich.edu? Many thanks!!
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How do you research and bring work together?
You may use the technique of consilience without knowing it.
Read this definition and then let me know how you use consilience in your work.
Highlights:
In science and history, consilience (also convergence of evidence or concordance of evidence) is the principle that evidence from independent, unrelated sources can "converge" on strong conclusions. That is, when multiple sources of evidence are in agreement, the conclusion can be very strong even when none of the individual sources of evidence is significantly so on its own. Most established scientific knowledge is supported by a convergence of evidence: if not, the evidence is comparatively weak, and there will not likely be a strong scientific consensus.
The principle is based on the unity of knowledge; measuring the same result by several different methods should lead to the same answer. For example, it should not matter whether one measures distances within the Giza pyramid complex by laser rangefinding, by satellite imaging, or with a meter stick – in all three cases, the answer should be approximately the same. For the same reason, different dating methods in geochronology should concur, a result in chemistry should not contradict a result in geology, etc.
The word consilience was originally coined as the phrase "consilience of inductions" by William Whewell (consilience refers to a "jumping together" of knowledge).[1][2] The word comes from Latin com- "together" and -siliens "jumping" (as in resilience).[3]
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Dear Colleague Michael Marek,
Yes, this is so often the case. My husband who is an active observational planetary scientist says how often the "devil" is in the details of data analysis.
Our short story collection, Children of Steel, is being considered by Wayne State UP, BTW. It is a collection of short fiction by people who grew up in steel mill towns. I just noticed where you retired from teaching.
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I am interested in Neurodegenerative diseases, particularly Dementia, and I would like to study the disease basic neurobiology. This will be a helping hand for me to study Dementia well.
Recommendations could be Books, Articles, Lectures, Animated videos..etc
Thanks in advance!
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Hi Mohab,
You could have a look at the NEURONET Knowledge Base
It provides a summary overview of the Innovative Medicines Initiative neurodegeneration research portfolio through an interactive dashboard, the NEURONET Knowledge Base includes links to over 500 publications and more than 380 publishable deliverable reports, acting as a one-stop shop to explore the diverse projects and outputs of the portfolio.
On the project website you can also find an overview of the ongoing (also completed) public-private research projects in the area of neurodegeneration (including dementia & Alzheimer's disease): https://www.imi-neuronet.org/ongoing-projects/
Hope this helps a bit.
Best wishes,
Chris
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lately, I've been thinking about whether or not mycelium can be used to link two brains or a set of neurons together. Mycelium is already known to be able to pass nutrients as well as electrical signals across the ground floor, so I theorize that it could be used to link neurons together or possibly even two brains. interested in what others have to think!
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Conversely, rotenone, which is an insecticide, has.
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in fly model MPTP can be used to induce parkinsons disease refer the following article for further clarification.
"Resveratrol prolongs lifespan and improves 1-methyl-4-phenyl1,2,3,6-tetrahydropyridine-induced oxidative damage and behavioural deficits in Drosophila melanogaster"
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Hello everyone!
We’ve started to work with this cell line, and it is driving us crazy. We are unable to making them attach to the plate and when we do, we see a huge amount of apoptosis and cell death.
The coating is performed with PDL (50 ug/mL) and laminin within a range of 6 to 12 ug/mL. Those parameters are what it’s written in most of the limited bibliography that exist about this cell line, so we are unable to find what’s the problem.
Has anyone experienced the same problem as us? Did you manage to solve it somehow?
The pics are showing how our cultures looks like in bright field microscope from 2 – 5 days after passage approx.
Thank you in advance!
Kind regards
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Dear Alejandro!
Please You look at the following protocol:
NSC-34 cells were cultured in differentiation medium consisting of minimum essential medium Eagle/alpha modification (Millipore-Sigma, Burlington, MA, USA) supplemented with 1% fetal bovine serum (Thermo-Fisher Scientific, Cambridge, MA, USA), 1% 100× MEM non-essential amino acid solution (Millipore-Sigma, Burlington, MA, USA), 1% pen strep (Thermo-Fisher Scientific, Cambridge, MA, USA). D
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Dear Colleagues,
Doctors from UCLA and Yale University are conducting a Survey on Postoperative Practices in Evaluating and Treating Patients with Brain Tumors in North America.
We are asking neurosurgeons, (neuro)psychologists, speech-language therapists, and occupational therapists, physiotherapists, or psychotherapists to participate in the survey.
Our goal is to understand common practices, disseminate standards of care, and gather information on post-operative outcomes in patients with brain tumors. We will publish the results from this survey in an open-access journal.
The survey can be accessed here:
Thank you very much for your help! Please reach out with any questions.
Monika Polczynska
UCLA Dept. of Psychiatry and Biobehavioral Sciences
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Shweta Singh Fantastic. Thank you very much. We are asking (neuro)psychologists, neurosurgeons, speech-language therapists, occupational therapists, physiotherapists, and psychotherapists to participate. If you have a few contacts you would like to share, please message me privately. We will be happy to reach out to these people directly, if it helps save your time. I am also providing my email address: MPolczynska@mendet.ucla.edu just in case. Best wishes, Monika
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I had a few students help me with a simple but time-consuming task. The data they helped with will be used in a scientific paper. The students are part of the Student Research Program (SRP) at my institution and they received a class credit for their work with me. Should I include these students as co-authors on the manuscript?
I also had a student volunteer help me on the same research project. The student did not get a class credit for their help. Should the student be a co-author on the paper?
Thank you in advance for your opinions/suggestions.
Monika Polczynska
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Co-authorship is not a form of reimbursement (in laboratories and research institutes, all co-authors receive a salary or grants, nevertheless...). If a persons has made a creative contributions, then they are a co-authors. If the work is technical - for example, reprinting a text, or measurements (without processing etc.), then - no. There is another form for highline such participation: acknowledgment.
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As i know , if we can realize the DNA structure , we can simulate it in computer . then we can try to rebuild it if possible .
so Given the technological progress, is it possible in the future?
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Compared to various living organisms characterized by a much lower level of organization and body structure, the human body has limited regenerative abilities. However, along with technological advances, in medicine, genetics, microbiological tests, etc., the possibilities of transplanting various organs, limbs, growing specific types of tissues and rebuilding certain parts of the human body are gradually increasing. One of the most difficult and perhaps impossible to implement in the future is the rebuilding of the central nervous system, including the human brain. Similarly, it will be extremely difficult in the future to build artificial awareness in artificial neural network systems as a continuation of the progress made in the development of artificial intelligence.
Best regards,
Dariusz Prokopowicz
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Please share your thoughts.
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When AI began in 1956 (Shannon McCarthy, “Autmata Studies”, 1956), there was no connection between AI and neurobiology. The two most influential people were Marvin Minsky and John McCarthy. Minsky said “solve the problem, ignore neurobiology”, while McCarthy believed everything could be solved by using formal logic. In 1956 the founders believed the problem of language would be solved in 10 years! (Still not, in 2021) In 1969 Minsky and Papert published a book where they proved that the perceptron could not learn to decide if a pattern was connected.
This effectively killed the neural network- like approach for the next thirteen years, until 1982 when John Hopfield showed that a recurrent network of 2-state neurons could implement content-addressable memory. AI became, roughly, theory of neural networks, influenced by neurobiology. Neuroscience continues to develop with essentially no influence from AI. Today, “deep learning” is the frontier of AI, with remarkable success, but its’ relationship to neuroscience is unclear. Today, it may be that the grid cell and place cells of neurobiology influence robotic spatial navigation. Phew! I hope this sketch is useful. On a personal level, as a graduate student in the 1970s, I was forbidden to use the words neuron or neural network in an AI thesis.
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Hello there, I am looking for this book to enrich my biological knowledge about the brain. Does anyone have it by any chance?
Also, would you recommend this book over the Kandel's "Principles of Neural Science" to examine the biology of the brain?
Thank you in advance,
Luca
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I would like to scan calcium-indicator-dye-loaded neurons after fixation, just to know, which cells have been loaded. Fura and Carbodiimide fixation have been reported to retain fluorescence but I prefer paraformaldehyde fixation for several reasons. Is there a single calcium dye, which is not quenched after paraformaldehyde fixation?
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May I ask if you continue your experiment and find a good agent? I am also thinking about fixing my cells on a coverglass right after treating with Fura-2 AM and I am wondering if fixation processes by 4% PFA may effect the membrane permeability and cause Ca2+ fluorescence to be lost.
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I need a stack of black and white .jpeg/.tiff/.jpg/.png images across time of either action potentials, neurons firing, or brain scans (comparing disease and normal brain, disease progression, etc.) that I can colorize and overlay for a project in a data visualization course. It wouldn't be published and only for submission to the instructor.
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Something like this for action potentials ?
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Dear Colleagues,
I am new to genes and genotyping and I need help. I am trying to figure our how AA, AG, and GG genotypes map on to A1 for DRD2/ANKK1 Taq1A polymorphism(rs1800497) and Val or MET for COMT Val158Metpolymorphism (rs4680). Where can I find this information presented in a way that a gene-naïve person will understand?
Thank you very much in advance,
Monika
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Hi Monika,
For me ensembl.org is the easiest. For example COMT
1. you search for the human gene
2. Open the gene file:
3. Select one of the longest and best annotated transcripts
4. Select cDNA in the left panel
If you don't see the SNPs go to Configure the Page and select Variants. If you prefer, you can visualize the SNPs as well in the gene view.
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Dear Colleagues,
Do you know of any applications that help adjust references in a manuscript to a journal's stylesheet? I am familiar with Mendeley but I was wondering whether there is program that does not require prior uploading cited papers (just as Mendeley does).
Thanks a lot!
Monika
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dear definitely many others apps there,but i suggest according user friendly convenience
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Dear Colleagues,
My team is planning to conduct a modified version of a scientific survey that was published by a different group a few years ago. We are going to significantly modify the survey and use it to investigate a different clinical population. We will, however, keep some of the questions used in the original survey. How should we best approach this without risking plagiarism?
We will say in future publications that will follow our survey that it is a modified version of a different survey. Which of the options below should we also pursuit:
(1) mention that our survey is a modified version of another survey already in the survey itself,
(2) paraphrase the questions that we will borrow from the original survey?
I will greatly appreciate your suggestions.
Thank you,
Monika Polczynska
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Request the pertinent permission from its Author or Authors and then make it reliable, validate it, etc. Pisometrically in its new modification and with its own "ad hoc" Normative Groups.
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I am doing research on the affect of shame.
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Similar to above. Jaak
Panksepp?
From affect/psychodynamic perspective Phil Mollon
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Let's talk about what is our self else than your memories (if all set of information that we've got is a different type of memories)?
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The self is a complex interaction between memory and attention. In this case memory is facts we know about ourselves (self-knowledge), schematic structures built from our socio-cultural context and autobiographical memory. The Self is how we appraise, use, and prioritize not only information in memory, but also various interests and desires. These actions on memory, interests, and desires require varying degrees of attentional resources. Carolyn Jennings makes this point much better than I in her essay (https://aeon.co/essays/what-is-the-self-if-not-that-which-pays-attention). In sum, there is a bidirectional relationship between our Self and how we appraise, use, and prioritize what’s stored away in memory structures as well as our management of our interests and desires.
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Does anyone have experience with the Olympus cellVivo weather chamber for monitoring neurite growth over hours/days?
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sorry not
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Hello,
I am trying to send triggers from MATLAB to the BIOPAC stimulator STM200. The aim is to deliver electric shocks to the participant. My STM200 is plugged into the STM100C through the 50 ohm output and the STM100C is placed in between the STP100 and the UIM100. The STP100 is connected to the stimuli presentation computer via a DB-25 ribbon cable.
Does anyone know how to send triggers in order to make the STM200 deliver a shock? I haven’t been able to find any example of code online.
Thank you in advance,
Chiara
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Andrew Chao no the project we were working on this for is currently stalled but will likely pick back up come May/June so if you figure this out it would be greatly appreciated. Likewise if we figure it out I'll send what ever we end up doing over your way.
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Dear Colleagues,
I am planning to introduce a new model in a paper. I have found a few good samples of published papers presenting new models in the area (neurolinguistics).
Are you familiar with any guidelines for research papers that introduce a model?
Thank you!
Monika
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Monika - the 'general' guidelines are similar to a 'primary' research paper in that the headings are similar i.e. Intro/background, methods (this may include an existing theoretical framework [as methodology] that you used to underpin the model and your literature search and appraisal strategy [if conducted]. It obviously doesn't include ethics, sampling, data collection/analysis.), limitations, further research, discussion, implications for practice, and conclusion/summary.
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What are your favorite and/or "gold standard" RNA-seq datasets for neurodegenerative diseases?
I come from the field of computational cancer genomics where there are some datasets that are well-known as "high-quality" ones in cancer. I want to benchmark some of the computational tools I'm currently developing on RNA-seq datasets for neurodegenerative diseases: I want to make sure I can identify known regulators with the tools I am testing. Therefore, I am looking for high-quality RNA-seq datasets for neurodegenerative diseases. Any advice on where to find such datasets?
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Hi Matvei Khoroshkin , the forerunner in RNA-seq data for AD and dementia and even TBI is the data atlas from Allen Institute which is quite solid and reliable and gives you many options to filter your results:
For PD RNA-seq data check:
You may also check the following website for papers with different RNA-seq data:
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 If so, what solvent did you use? What was the max concentration of cuprizone you used to make the stock solution? I am having trouble replicating a protocol in a paper that made a 1mM cuprizone stock in 50% ethanol, then used the cuprizone on cells at a concentration of 500uM. This just causes a precipitate likely due to the high ethanol concentration (25%)!?
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I was also asking myself your question and then I found
not sure if it is still important to you but it may be for others...
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The feeding habit and the psychological disorders.
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Please take a look at the following PDF attachments.
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Most of biological and medical investigations choose to involve either male or female (rarely both) subjects to study.
To merely include subjects from one gender (particularly males in rodent models in many studies) and to avoid populations of mixed genders in separate groups is almost a very well-established approach in experimental design of most of the studies in neurosciences, brain and cognitive sciences, cancer research, immunology and behavioral psychology, etc.
Nevertheless, I think that such an approach is often disputatious and may be inaccurate (though statistically solid only for those experimental groups), in that the real world population is not segregated, and research should truly reflect on the statistically unbiased real demographics.
Do you think that to avoid statistically unreliable variances due to biological gender differences in a mixed study (not mixed-gender groups), and thus to choose subjects from only one gender (male or female) would result in biased, unreliable or non-replicable studies?
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It is only when you are working with rodents you have to abstain from using females because their estrus cycle disturbs the results.
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I'm unfamiliar with the contemporary technology in neurobiology. I'm interested in the possibilities to measure very small energy fluctuations in the brain, the location of the activities is rather unimportant for my purpose. I would be very grateful for any suggestions regarding the relevant literature.
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Hello I am not an expert on that so pardon but it would seem to be on the order of one or a few photons depending on your theory and what fluctuations you speculate matter. Either you are looking for quantum level changes which you could capture with a total surround of the brain or a partial surround with compensatory assumptions, or you are looking for much more macroscopic change such as heat flow, where the resolution would of course be rather coarse particularly with probable difficulties arranging for a tightly controlled experiment.
Put another way, with less confidence you can make assumptions that make the test easier and at least reduce the hypothesis set thereby.
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Dear all,
For the analysis of intracranial electrophysiological recording in animals, some tutorials montion that the analysis should be used for stationary signal, so there should be some preprocessing, like prewhitening. 
Some tutorials also said some signal could be thought as locally stationary. So I am puzzled, should I use prewhitening, and when should I do this?
Particularly, the analysing procedure is like this: reading the raw data, then do low-pass filter for LFP, then analying power spectrum of LFP, etc. High-pass filter for spike, then spike-sorting, then estimating firing rate, then do some statistic analysis
By the way, I am using Matlab toolbox, like Chronux or Fieldtrip.
Best wishes,
Jichen
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Oh, the phase-locking between spikes and theta LFP change with the moving speed. I get it, thank you!
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Neuro 2A (N2a) is a mouse neural crest-derived cell line that has been extensively used to study neuronal differentiation, axonal growth and signaling pathways. Is it also possible to record action potential from these cells? Any description, idea and suggestion will be appreciated.
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Can one use N2a cells s.c. injection to study tumor growth in other mice strain then AJ?!
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Could you imagine to create a video explaining the essential of you publications yourself?
Why would you want to create a video for you publication?
Did you produce these kind of Videos already? If so, how did you do it?
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Journal of Visualized Experiments (J Vis Exp, JoVE)
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I would like to transduce neuronal and astrogilal primary culture cells from mice or rats with an AAV.  Do you know a reliable article that allows to answer to these questions :  Should I produce an AAV2/6 ? Should I transduce at DIV6 or DIV2 ? 
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Hi Marie
You could try AAV-DJ, which is the most efficient serotype in infecting in vitro cells in our experience. Paradoxically, AAV-DJ is not good in tissue infection in vivo.
Genemedi is experienced in AAV production, you could find more information and manual files on this website: www.genemedi.net/i/aav-packaging
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Dear Colleagues,
Are you familiar with any studies on the amount of language switching (code switching, language mixing , or both) on the organization of languages in the brain?
Thank you!
Monika
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And here are more titles:
Schwartz, M. (1994). Ictal language shift in a polyglot. Journal of Neurology, Neurosurgery and Psychiatry, 57, 121.
Javier, R.A. & Marcos, L.R. (1989). The role of stress on the language-independence and code-switching phenomena. Journal of Psycholinguistic Research, 18(5), 449-472
Green, D.W. (1986). Control, activation and resource: A framework and a model for the control of speech in bilinguals. Brain and Language, 27, 210-223
and of course
Albert, M.L. & Obler, L.K. (1978). The Bilingual Brain: Neuropsychological and neurolinguistic aspects of bilingualism. New York: Academic Press
There are so many other, just let me know if you need more. Good luck!
Christina
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Dear all,
Which medium to use after differentiation of SHSY5Y? I used RA+BDNF and I need to use different treatment for next 2 days. Keep them in the differentiation medium or I can use standard medium for SHSY5Y cell excl. FCS, or they need some supplements?
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Tanisha Singh,
Thank you so much for your answer and advice.
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Does anyone know how/if its possible to measure erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA) in living humans?
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آسف السؤال ليس من اختصاصي
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Is there any software for drawing synapses and neurons?
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BioRender might be a good option (https://biorender.io/)
It's an online app that contains a library of pre-made cells, proteins, membrane shapes, organs, lab equipment, etc. that you can drag-and-drop so you don't have to spend time drawing each element of the figure yourself. Saves a LOT of time and the final figures you create are professional-looking. Plus, it's free for educational use.
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DNA structure and activity can change with environmental factors. But does experience such as sexual abuse create significant biological effects on DNA?
These type of researches are so important in detecting crime.
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Just like environmental factors, psychological factors may have an impact on gene expression and its regulatory mechanisms but the extent of impact would be depndent on multifactorial aspects and would need to be ascertained in a case to case basis.
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Dear Colleagues,
Are cognitive control and cognitive effort the same thing?
I am confused. In the neuroscientific literature on second language learning, the neural organization of cognitive control and cognitive effort seems to overlap (e.g., the anterior cingulate, dorsolateral prefrontal cortex, inferior parietal regions). At the same time, I have come across some results that make me think that cognitive control and cognitive effort are not the same thing.
For instance, I have found fMRI studies that reported increased "cognitive effort" (and therefore more widespread fMRI activation) in less proficient speakers of a second language (e.g., Abutalebi et al. 2018). At the same time, several ERP studies have suggested more "cognitive/language control" in highly proficient second language users. For example, Fernandez et al. (2013) have shown that higher second language proficiency was associated with a greater mean N2 amplitude (greater inhibition) on an executive function test. Another example: Rossi et al (2018) concluded that individuals with high second language proficiency require more cognitive/language control for their first language, even before they speak their second language.
I will greatly appreciate your help.
With my best wishes,
Monika
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Dear Colleagues,
Are you familiar with neuroimaging studies or/and do you have any predictions about executive control in implicit/informal versus explicit/formal second language learning? Which type of language learning requires more executive control?
I will greatly appreciate your opinions/suggestions.
All the best,
Monika
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Dear Monika,
Results in some recent meta-analyses (e.g., by Shaofeng Li) have shown that executive control plays a more significant role in explicit/formal than in implicit/naturalistic L2 learning (SLA). Regarding implicit and explicit brain networks, a similar pattern can be detected, e.g., in the paper by Jing Yang & Ping Li (2012). Brain networks of explicit and implicit learning. PloS One, 7(8), e42993.
Hope this above information helps a bit,
Best,
Edward
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Corpus callosum is the main bond between the brain’s left and right hemispheres. It is sometimes damaged or cut out because of a disease, this is called split brain, in this case the two halves of brain do not exchange information efficiently. In one experiment a word was flashed briefly to the right field of view of a split brain patient and the patient was asked what he saw and because the input from the right field of view is processed by the left hemisphere which is also responsible for verbal processing the patient’s answer matched the word, next the word was flashed to his left field of view and the experiment was repeated and the patient said that he did not see anything but when he was asked to draw a picture with his left hand he drew a picture of the word. The question is are there some stimuli in our environment that we can’t sense but affect our decisions? In other words do we decide or our behavior may be a reaction to the things that we are not aware of?
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It seems similar to what is told to happen in hypnosis: telling a person to do something after hypnosis, that person ( sometime) can do it after hypnosis without remembering the order.
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We are a team including Epidemiologists, Physicians, Biologists (MSC and PhD), Well-known supervisors from Iran, United States and India and we are writing a lot of articles in these countries.
Now, we need more authors in our team. Motivated, active, Smart and clever authors from everywhere in the world.
Currently we want to form a team and write a new review article in Neuroscience (Especially on Impacts of Nutrients on Brain developments).
Kindly send your request by E-mail or direct message on RG.
Do not hesitate to ask any question.
Best of Luck
Kheirvari JK
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Good Luck
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We want to characterise some brain regions in an insect using an anti-synapsin protein. Most protocols use Normal Goat Serum to dilute the antibodies and as a blocking solution, but it seems to be difficult to get here in Mexico. There are alternatives, like BlockAid, a proprietary formula that is better at blocking unspecific binding (according to the manufacturer). The problem is that I haven't found a single paper where BlockAid is used in insects with our antibodies. Has anyone used these solutions? Are they really better? What are the possible drawbacks?
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There are a number of considerations to take into account when using your blocking reagent. Using whole serum looks attractive on paper as it contains several proteins, yet the major protein is serum albumen. If you use whole serum, there will also be IgG's in there, and it is for this reason that people use whole serum from the same species that was used to raise the secondary antisera [think what would happen if you were to use the whole serum from the same species in which your primary antisera were raised]. In the past I have used secondary pig antibodies, and normal pig serum was easily obtained from an abattoir. Most of my secondary antisera are made in goats and I use normal goat serum. For many antisera BSA will work just as well, however I never try that as I have many neuropeptide antisera in which BSA is used as the carrier protein and hence my primary antisera may contains anti-BSA IgGs. If anti-BSA IgG's were to become a problem other blocking reagents like the one suggested here (fish skin gelatin) or non-fat milk powder may work as well.
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I am trying to design carbonate apatite nanoparticles that might cross the endothelial layer of brain capillary (blood brain barrier-BBB). Thus, an in vitro system would be a better option (easy to understand and faster) to check the permeability of particles before moving to an in vivo model. I am looking for an in vitro BBB system that can be used for this purpose. I have seen the co-culturing systems discussed by many researchers. Are the co-culturing system with the filter available commercially?
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You can look up Mimetas and easily make a BBB model with their OrganoPlate https://mimetas.com/overview-mimetas-applications/human-blood-brain-barrier
There was a paper published in February this year in Scientific Reports with this platform: https://www.nature.com/articles/s41598-018-20876-2
Hope it helps!
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In a situation where a myelinated neuron (at any location in the nervous system) gets demyelinated, what are the changes that occur in the neuron. May be there are three phasic changes. Pre-Demyelination changes, Intra-Demyelination changes and Post-Demyelination changes. What are these changes? Can we ennumerate them.
Important: Does any change in the size/length of the neuron occur at any given instant during the process of demyelination?
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Look, it's a good question ... I try to give a ontological answer but also (above all) a functional one and, in the end, it is the applied chemical stoichiometry that will tell us if budgets exist that do not return and in those mysterious areas we must go to investigate. Chemistry will not support the needs of dialectics even if expressed by authoritative scientists. Truth wins in science, and it is evident that chemical stoichiometry makes this research more fruitful.
The cell body of the neuron needs a lot of energy and the mitochondria can not provide everything that is needed. However, the mitochondria contained in the cell body are very few and this is just a conundrum: in fact it is paradoxical that a cell that consumes a lot of energy with a request for high oxygen supply for biological combustion has few mitochondria!
Different facts lead us to think that, thanks to the probable fusion of the mitochondria with the Nissl substance (Nissl: the endoplasmic reticulum of the neuron-soma) this gives the soma the “machine” that synthesizes all the ATP it needs but… in the very long axon that happens? The endoplasmic reticulum does not penetrates the axon and it’s true-like that myelin support this energy demand, especially for firing (see " Monitoring ATP dynamics in electrically active white matter tracts” Elife. 2017 - PMID: 28414271) let us not forget, is energetically dispersive. So the white matter, but also the gray matter, energetically supports the axon with myelin. Among other things, there is evidence that neural activity stimulates myelin growth. Keep in mind that the brain has a serious deficit, compared for example to the muscle: it does not possess myoglobin which is slow reserve of oxygen so much that it is extremely affected by ischemia, while for the muscle this criticality does not exist, thanks to myoglobin.
So myelin is home of an active energy metabolism with aerobic degradation of glucose, which, through the Krebs cycle - which we have shown to be very active in myelin (see our article “Tricarboxylic acid cycle-sustained oxidative phosphorylation in isolated myelin vesicles” PMID: 23851157), supports "outside" the neuron's energy needs, above all to support firing.
But there's more!
In fact, for go on the combustion the myelin need a lot of oxygen and that's why the myelin has the highest content of carbonic anhydrase (CA) of the whole organism. Let us remember that CA is the enzyme that combines the carbon dioxide produced in gaseous form with water to make it soluble in the form of bicarbonate. Remember us that if the CO2 remained gaseous (let's remember that in this state it is produced by the Krebs cycle) in the brain we would have terrible seizures! Furthermore, again to be in the subject of oxygen, myelin dissolves oxygen quite well and therefore constitutes a slow reserve of oxygen to support neuronal combustion, even if it is not as efficient as myoglobin. In fact, white matter resists better the hypoxia of gray matter.
So many data would then remain mysterious if we do not take into consideration the chemical reactions we have now mentioned are operative in myelin.
At last to answer with the ontological profile, I hope that I have answered your question, and we can realize that all this chemical support to the axon from myelin could not be supported by the neuronal cell alone. I believe that it is possible to look at the issue from a teleological point of view, but if we talk about it we will talk about it another time…
Yours sincerely,
Sandro
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In several papers I found that testosterone might inhibit oxytocinergic activity in the brain, but no references were given.
For instance, I already read that oxytocin makes the orbitofrontal cortex more active (in some tasks) shifting away the activation from the amygdala, while testosterone does the reverse activating the amygdala and “deactivating” the OFC. But there is no mention on how these hormones interact.
In other papers, I found that testosterone inhibits oxytocin, but no references are given. For example: “we chose female participants in luteal phase because testosterone might interact with endogenous oxytocin”.
So, I’m asking whether this is a general and accepted chemical phenomenon “testosterone over oxytocin”, or rather if this depends on singular cases.
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Ok, after two years, I can come up with a partial answer!
Testosterone (T) and oxytocin (OXT) are well known to interact with each other (1–3), producing opposite behavioral effects (4–7) due to the blocked OXT receptors by T (8,9). In the brain, one of the target of both T and OXT is the prefrontal cortex (PFC) (10), a network that is crucial for impulse control and self-regulation, important also for social cognition, mentalization, and empathy (11). The PFC has an inhibitor effect on behavior (12,13), thus, a reduced activity does lead to an increased propensity towards aggressive behavior (14).
References
1. Gossen A, Hahn A, Westphal L, Prinz S, Schultz RT, Gründer G, et al. Oxytocin plasma concentrations after single intranasal oxytocin administration - A study in healthy men. Neuropeptides [Internet]. Elsevier Ltd; 2012;46(5):211–5. Available from: http://dx.doi.org/10.1016/j.npep.2012.07.001
2. Dhakar MB, Stevenson EL, Caldwell HK. Oxytocin, vasopressin, and their interplay with gonadal steroids. In: Oxytocin, Vasopressin and Related Peptides in the Regulation of Behavior [Internet]. 2013. p. 3–26. Available from: http://ebooks.cambridge.org/ref/id/CBO9781139017855
3. Frayne J, Nicholson HD. Effect of oxytocin on testosterone production by isolated rat Leydig cells is mediated via a specific oxytocin receptor. Biol Reprod [Internet]. 1995;52(6):1268–73. Available from: http://www.ncbi.nlm.nih.gov/pubmed/7632835
4. Burnham TC. High-testosterone men reject low ultimatum game offers. Proc Biol Sci [Internet]. 2007;274(1623):2327–30. Available from: http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=1950304&tool=pmcentrez&rendertype=abstract
5. Zak PJ, Kurzban R, Ahmadi S, Swerdloff RS, Park J, Efremidze L, et al. Testosterone Administration Decreases Generosity in the Ultimatum Game. Aleman A, editor. PLoS One [Internet]. 2009 Dec 16;4(12):e8330. Available from: http://dx.plos.org/10.1371/journal.pone.0008330
6. Zilioli S, Ponzi D, Henry A, Maestripieri D. Testosterone, Cortisol and Empathy: Evidence for the Dual-Hormone Hypothesis. Adapt Hum Behav Physiol [Internet]. 2015;1(4):421–33. Available from: http://link.springer.com/10.1007/s40750-014-0017-x
7. Zak PJ, Borja K, Matzner WT, Kurzban R. The Neuroeconomics of Distrust: Sex Differences in Behavior and Physiology. Am Econ Rev [Internet]. 2005 Apr;95(2):360–3. Available from: http://www.jstor.org/stable/10.2307/4132847%5Cnpapers3://publication/uuid/BB68F281-21C7-452C-9238-C59CBE7560DA
8. Insel TR, Young LJ, Witt DM, Crews D. Gonadal steroids have paradoxical effects on brain oxytocin receptors. J Neuroendocrinol [Internet]. 1993;5(6):619–28. Available from: http://www.ncbi.nlm.nih.gov/pubmed/8680433
9. Arsenijevic Y, Tribollet E. Region-specific effect of testosterone on oxytocin receptor binding in the brain of the aged rat. Brain Res. 1998;785(1):167–70.
10. Bos PA, Hofman D, Hermans EJ, Montoya ER, Baron-Cohen S, van Honk J. Testosterone reduces functional connectivity during the ‘Reading the Mind in the Eyes’ Test. Psychoneuroendocrinology [Internet]. Elsevier Ltd; 2016;68(March):194–201. Available from: http://linkinghub.elsevier.com/retrieve/pii/S0306453016300671
11. Frith CD, Frith U. The Neural Basis of Mentalizing. Neuron [Internet]. 2006;50(4):531–4. Available from: http://linkinghub.elsevier.com/retrieve/pii/S0896627306003448
12. Goldstein JM, Jerram M, Poldrack R, Anagnoson R, Breiter HC, Makris N, et al. Sex differences in prefrontal cortical brain activity during fMRI of auditory verbal working memory. Neuropsychology [Internet]. 2005;19(4):509–19. Available from: https://www.tandfonline.com/doi/full/10.1080/10253890.2017.1378638
13. Güvendir E. Why are males inclined to use strong swear words more than females? An evolutionary explanation based on male intergroup aggressiveness. Lang Sci [Internet]. Elsevier Ltd; 2015;(13):1–7. Available from: http://linkinghub.elsevier.com/retrieve/pii/S0388000115000194
14. Mehta PH, Beer J. Neural Mechanisms of the Testosterone-Aggression Relation:The Role of Orbitofrontal Cortex. J Cogn Neurosci [Internet]. 2010;22(10):2357–68. Available from: http://www.ncbi.nlm.nih.gov/pubmed/19925198
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I have TTX citrate ordered from Hello Bio. On using it at 1uM as my working concentration, it didn't show any effect.
How do I test whether the toxin is actually working or not?
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Dear Yojet,
I suppose you have patch clamp rig to measure ionic currents, if yes, then you can simply patch NSC derived neuron and record inward INa and apply TTX. You shall see rapid and almost complete block of sodium currents (see attached pic) which will confirm the efficacy and potency of TTX stocks you have. You can refer to the following paper for internal external combination and protocol to be used for recording.
Ensure that you aliquote your TTX and store it at -20/80 to preserves its efficacy. I hope it helps!
Best,
Nand
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I plan to use Cogent for the event-related presentation of my stimuli. The stimuli are short video clips (3 seconds) and I have to decide in which format the videos should be. I can convert them to any format possible. At the moment the videos are in .wmv format, but I heard that this format may not be supported in Cogent. Is this correct? Is there also a video codec specified that is supported in Cogent?
At the moment I cannot retrieve the website of the Wellcome Laboratory of Neurobiology and I do not know why (there is always a error message). For this reason I cannot check the information by myself or download the Cogent toolbox to test it.
Thank you very much for your help!
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Hi Lorena
Videos should be in .avi format. If you send me a mail I can share the toolbox with you.
Best
Julian
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Cells not easy to transfect
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Did you ever try transfecting primary motor neurons with AMAXA?
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Hi all,
I recently begin to learn single-unit recording in the primary visual cortex of mouse brain using tungsten electrodes. Sometimes (but not very often), there is significant bleeding during when I try to remove the skull and the meninges, but in all the cases the vessels eventually stopped to bleed. I wish to know how will bleeding affect neurons nearby? Will neurons die because they have less O2 and glucose supplied?
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Dear Zhou,
That will depend on the level of tissue and bleeding. If there is significant level of blood supply to the area that you are recording from, the reduction in local oxygen supply can cell death. Another factor is the swelling associated with damage to the Pia mater and damage other tissue ( interstitial) damage. Usually as you go down with your electrode you should be able to see/hear (from amplifier) discharges of damaged neurons. If your electrode tip is not blocked and you hear/see no discharges at all. That is bad news, meaning substantial damage. That is what I used to do in my studies, for an example see below.
Best wishes,
Refik
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Greetings,
While PC-12 cells are commonly differentiated using nerve growth factor (NGF), I am going to be using PC-12 Adh (the adherent version of PC-12 cells) for a series of experiments involving NGF.
Here are my questions in case you have used NGF on PC-12 Adh cells:
- What media/supplements did you use for maintenance as well as when differentiating the cells?
- Did you find that the cells grew better in certain plates/flasks?
- What markers did you use to confirm differentiation? Neurite growth/length, gene expression, protein levels, etc.?
Thank you for your help!
Shahin
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Just to avoid you a big waste of time; PC-12 Adh do not differentiate/grow neurites. I tried every possible experimental setups. They are not sensitive to NGF.
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Fluorescent dye in uncoupled astrocytes,  and Calcium activity. 
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I dissolved the carbenoxolone disodium salt in distilled water. However, after mixed the carbenoxolone with saline solution, there is a lot of cloudy things. Does any one see this phynomena?
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Hi. Recently I've tried to record field potential in brain slice but failed. I use bipolar or monopolar stimulating electrode. Amp is Axopatch 1-D and headstage is CV-4. Recording was done at I-clamp mode. When the recording electrode containing normal ACSF touched the surface of brain slice, I started current injection but I could not see any responses but stimulus artifact. Would you please give me some advice?
As I had not recorded field potential before, I used brain stem slice (I have many experiences here). Age of mice is around P(postnatal day)3~P9. It is MNTB-LSO synapses at pons level.
Stimulating electrode(bipolar or unipolar) was located at MNTB and recording electrode (3~5 megaohm) was at LSO. The resistance of stimulating electrode was also in the same range of that of recording electrode (in case of unipolar electrode). I also tried bipolar electrode but failed.
LSO cells viewed with high magnification were alive and healthy. In voltage clamp mode, postsynaptic currents were elicited by stimulation at MNTB.
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Hi Seungcheol Ahn,
Thanks for the update. That is a non-trivial slice, and one that I don't have personal experience with. My worry would be that you have cut the axons between the MNTB and the LSO.
If I were you, I would cut hippocampal slices. Pretty much any slice you cut that has a complete hippocampus in it will have a connection between CA3 and CA1. If you stimulate in stratum radiatum and put a recording electrode in the pyramidal cell layer you should see a large population spike (> 1mV). If you can not see this, then there is a problem with your stimulating or recording system.
However, if I had to make a guess, other than the worry that you have cut the MNTB LSO axons, it might be that the resistance of your stimulating electrode is too high. If we say that it has a resistance of 5MOhm, and you want to pass 100uA of stimulating current, then that would require 500 Volts, which is far more than any stimulating system that I know of has available. You'd be limited to less than 20 uA, which is starting to get pretty low (not that that amount can't work, it's just not what I would recommend for testing a system).
Try stimulating with a simple Tungsten wire (or something similar), with a resistance of 100kOhm or so.
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Dear colleague,
I would like to stimulate my recorded cell (whole-cell, pyramidal cells in hippocampus) with a pulse train of light during 600 ms. In the middle of those pulses of light, I would like to stimulate electrically the axons (0.1 ms, 1 pulse). How can we "superimposed " the two stimulations with clampex 10 ? For now I found how to build the protocole with one stim after the other. But I would like to stimulate electrically while the pulse of light is ON.
My electric stimulator is branched on the Out#0 and the LED is branched on the Out#1 of a Digidata 1500A.
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Under the waveform tab you set up three waveform epochs, the first one is just light, the second one is light plus stimulus and the third one is just light again.
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For the axon morphology analysis, I found that it is very difficult to get a clear conclusion with hippocampal neuron culture, because there are lots of varieties exist with hip neuron. I am wondering that whether this is reason people prefer to choosing spinal neuron for their axon morphology study. Just like"Dynamic Localization of G-Actin during Membrane Protrusion in Neuronal Motility" also use Xenopus Spinal Neuron Culture.
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Vlad, probably the question is the sentence right after "I am wondering (...) whether...". Isn't it?   :v
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Hello,
I'm trying to image neurons every 15 min for 24 hours. My neurons don't even get past the 6h mark before dying. 
I plate my neurons on precoated ibidi chambers and use Dil at 5ul/1ml from 1mM stock with an incubation time of 5 minutes. After labeling, I replenish with fresh media plus some of the old media (growth factors) with added FBS. I've been adding 10% FBS because a neighboring lab found that longterm imaging without FBS of labeled neurons can cause the neurons to die.
The causes of death I'd like to rule out are (1) FBS or (2) Dil concentration/incubation time. Previously I had been imaging via phase contrast with phenol red in my media and that didn't seem to harm the neurons (although this was a 12 hr time lapse). 
If anyone can help me out I'd greatly appreciate it!! Thanks!
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 Hi Angelica,
Depending on the specifics of your experiment, you might also try doing transient transfection of a membrane bound fluorophore (e.g. mGFP https://www.addgene.org/14757/). I've done such transfections in on primary neural culture using CaPO4 method and Lipofectamine. In each case, the key was to optimize concentrations and incubation.
Best of luck, 
Joey 
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Looking at neurobiologic justifications and findings in education, for example music education: What can you really tell about this field's research results that has not already be known before?
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Please check the material at the link I sent you on the previus message (this one: http://cienciaparaeducacao.org/eng/publications/)
Best,
Mateus
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We have homogenized Taenia crassiceps larvae and are puffing the homogenate onto pyramidal neurons during whole-cell patch-clamp recordings in organotypic hippocampal slice cultures. We need to put the puffer pipette very close to the neurons - ie the neurons move during the puffing - we see obvious depolarization ie 10 - 20 mV worth, that is not blocked by glutamate receptor blockers (kynurenic acid, AP5, CNQX). The pH is roughly between 7 and 8. Osmolarity of the homogenate is 300 ish. Also the K+ concentration within the homogenate is 4 mM and the effect is there when using a caesium internal. Is what we are seeing  an artefact?  What substances cause neurons to depolarize, what should we be thinking of as causative agents that might be in the homogenate?
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Most likely the homogenate contains glutamate as it is near millimolar concentrations in cytoplasm which will activate glutamatergic receptors in the pyramidal neurons. Glycine concentration is also likely sufficient to activate GluN1 subunits of heteromeric NMDA receptors. I suggest coapplication of ionotropic glutamate receptor antagonists, eg. CNQX and APV.
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Thank you for your kind in advance.
I am cultivating neural stem cells from E14 rat spinal cord and plate them into 100mm dish by dissociating accumax or accutase.
I  want to maintain neurospheres, not attaching cells, which might be differentiated over time. That`s why I don`t want to attach neurospheres until I want. I always plate them into 100mm dish that not coated anything for cell culture. The neurospheres are dissociated mechanically by pipetting up and down. and the neurospheres are always fine until 1week I dissociate it.
But, After 1week, I usually dissociated neurospheres into sing cells.
At the same procedure as above, I dissociated it and plate it into 100mm dish again. but, it always initiate to attach at the bottom of plate. There were not a lot of free floating cells as single cells.
I am facing with this problem.. Why do neurospheres start to attach at the bottom of plate? even though all of procedure was same, 2nd passage neurospheres initiate to attach bottom? anyone help me?
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Hee-Hwan,
In my experience with brain derived neural stem cells, there are a number of things that affect the maintenance of the the stem cells and the development of the neurospheres. 
Cell passage: how high is the passage number of your cell line? The higher it is the less likely the cells will behave the way you want.
Expansion time: you say your are passaging the cells every 7 days or so.  How big are the neurospheres at this time?  the smaller the spheres, the better.  It is when the neurospheres get really big (1-2 millimeters in diameter) that the cells begin to differentiate.  When this happens and you dissociate the spheres, stem cells which have started differentiating down a glial cell fate will attach to the bottom of the plate.  The floating cells are either dead cells OR are still undifferentiated. When there are only a few of these floater cells, it might be better to re-plate them in a smaller dish, then expand them into a larger dish in 2-3 days (do not passage them, just dissociate when they are 10-15 cell spheres and put everything back into a larger dish).
Serum and growth factors:  Sometimes serum contains growth factors that encourage differentiation.  It is better to use a serum-free media and add growth factors that help maintain the undifferentiated state (ie. bFGF). 
It is good that you are not coating your dishes.  You might even try non-tissue culture dishes, low attachment dishes, or bacterial dishes to reduce attachment.
I've listed some good references below, although they are all brain based neural stem cells.  It is possible that spinal cord stem cells behave entirely differently.  What kind of cells are you hoping to get? glia? schwann? neurons? Gage's seminal paper in Science is a good place to start although keep in mind the field has advanced a lot since his original paper 17 years ago. 
Hope this helps!  Cheers, Kim
Gage FH. Mammalian neural stem cells. Science. 2000 Feb 25;287(5457):1433-8.
Review. PubMed PMID: 10688783
Rietze RL, Reynolds BA. Neural stem cell isolation and characterization.
Methods Enzymol. 2006;419:3-23. Review. PubMed PMID: 17141049.
Marchenko S, Flanagan L. Passaging human neural stem cells. J Vis Exp.
2007;(7):263. doi: 10.3791/263. Epub 2007 Aug 22. PubMed PMID: 18989434;
Kim HT, Kim IS, Lee IS, Lee JP, Snyder EY, Park KI. Human neurospheres derived
from the fetal central nervous system are regionally and temporally specified but
are not committed. Exp Neurol. 2006 May;199(1):222-35. Epub 2006 May 22. PubMed PMID: 16714017.
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I have noticed an increased in the asymmetrical synaptic excitatory junctions to be increased in the diabetic brain but could not find any discussion of this in the literature at present.  If you have any information or publications regarding this question please share with me when you have the time.  
My hypothesis is that in younger diabetic brains this increase in asymmetrical synaptic excitatory junctions may result in an overuse phenomenon and result in a later or older models in their attentuation and or loss as in accelerated aging in the diabetic brain.  Thank you for your time and consideration.
Sincerely,
M.R. (Pete) Hayden, MD 
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Dr. S. Liu,
Your answer was very much appreciated and I have learned a great deal due to your paper in PNAS.  I am going to be studying the db/db K strain model in the coming months, while I will be focusing my TEM  studies of this model primarily on the neurovascular unit I will also be evaluating the  dysmorphic myelin white matter and due to the better understancing of you PNAS paper I will be keeping a close observation regarding the assymettrical excitatory snapses as well.  You paper really helped and I am so thankful for your earlier response to my questions regarding an observation of increased excitatory synapses in the type 1 model and now will be alert to observe similar changes in the obese T2DM of obesity insulin resistance hyperinsulinemia  and persisitent elevation of glucose levels well above the diabetic range for mice  - approximately 300-400 ranges. 
Sincerely ,
M.R.(Pete) Hayden MD 
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I am using ALexa 594 Fluoroscent dye. In acute hippocampal slices from adult rats, I fill the CA1 pyramidal neurons with the dye after getting whole cell configuration. I perform uncaging experiments for over 30 mins and wish to calculate and report the structural changes in the dendritic spines that I am uncaging on. Using 2-photon, I take z-stacks of the dendrite I am uncaging on and also movie of the dendrite. Can someone please tell me the best way/protocol to analyze the sturctural offiline (ImageJ, MATLAB)?  
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Dear Antant,
as a beginner in image analysis I found this book extremely helpful, practical and also entertaining:
I also have experience with matlab. Matlab can be connected with ImageJ and fiji:
So you can combine the two softwares. I like that, because I can immedately create the graphs as well. 
Good luck with your analysis!
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This question is specifically asked within the Project called "Treatment of Melanoma Brain Metastases."
Because neurological tissues as well as many forms of skin malignancies will tend to express at least some significant CB1 and CB2 receptors, perhaps using moderate dosing of a safe agonist like Delta 9-THC may be useful?  And consider including evaluation of equivalent to 100-300 mg or oral CBD for humans as well as this is becoming very common among patients.  
I work with both brain cancer patients (mostly Glioblastoma) and many breast cancer patients and they had already chosen to integrate cannabis extracts into their therapies. Because CBD and THC cross the BBB, and does not appear toxic to healthy normal cells, it may be reasonable to consider exploring this with research. I do have one interesting patient who reported using these compounds to treat her ER-PR-HER2+ brain metastases with tremendous success in only 3 months. The Herceptin and Perjeta she was on are too large to cross the BBB, so the situation is dire for her otherwise. Best wishes and thank you for your project!
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OK.  I think it is a good idea.
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Can you use 4-AP to assess excitability in dendritic FIELD POTENTIAL recordings from medial perforant path to dentate granule cell pathway?
I am trying to assess excitability of MPP-DGC (medial perforant path-dentate granule cell) synapses and have 4-AP available in lab. I am already doing other field experiments animals and therefore cannot do whole-cell yet to answer this question. I was wondering if I could get some prelim data in the mean-time by getting a baseline field recording at MPP-DGG (1 stim, every 30 sec, 100us duration) and then apply 4-AP (100 uM) to assess changes in excitability when Kv channels are blocked. 
Just need to know if I am even in the ballpark for assessing excitability at these synapses in each group. 
I am already doing separate experiments with picrotoxin and therefore am already looking at inhibitory transmission in this regard. 
Please help!
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Thank you for your timely response William! This does help a lot. Information about how well the cell can get signals to the the soma is still pertinent information in my study. 
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What is the role Calcium in Neuro-degenerative disease,… especially PD,AD,ALS ect whether its increases or decreases,as there is variant data available in the literature,...such as in Intra-cellular Calcium,Extra-cellular Calcium,Neuronal Calcium, Serum Calcium,Cytosolic Calcium and Mitochondrial Calcium?
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In neurons, calcium entry is performed as Dr Refik Kanjhan say.  We only wan to indicate that this entry comport a high intracellular Ca2+ concentration which it is dangerous by the cells. Normal cells remove Ca2+ as Dr Refik Kanjhan say. However, in the case of neurodegenerative disease, cells are damaged and are unable to use these mechanisms to removed intracellular Ca2+. Consequently, this Ca2+ may induce: 1) activation of protolytic and lipolytic proteins as calpaines and so on, 2) dysfunction of mitochondria.
Between  calpaines functions are the inhibition of Na+/Ca2+ transport, the most implied in remove de intracellular Ca2+.
The mitochondria dysfunction involves mitochondrial membrane depolarization and cytochrome c release. The cytochrome c release induces caspase 9 activaton, this caspase 3 activation, this nuclear degradation and apoptosis and cellular death.
Mitochondrial dysfunction also induces ROS formation, this causes lipid peroxidation and cellular death by necrosis. In neurodegenerative diseases the ROS are increased
Ca2+ may also activate some genes with the consequently dysfunctions.
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Many groups are nowadays trying to stimulate nervous tissue electrically or optically to achieve some intended therapeutical results. 
However, the usual staining methods for microglia or astrocyte activation  seem at odds with a short term overstimulation (kind of single exposure...). In acute settings, the body doesn't have the time to activate these pathes....
Does anybody have a recommendation how to quantify nerve or tissue damage below the threshold of literally "cooking" or "carbonizing" tissue? (That would be easy to recognize, though...) 
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Wells, J. D., S. Thomsen, P. Whitaker, E. D. Jansen, C. C. Kao, P. E. Konrad and A. Mahadevan-Jansen (2007). "Optically mediated nerve stimulation: Identification of injury thresholds." Lasers Surg Med 39(6): 513-526.
uses "hematoxylin and eosin (H&E) to demonstrate pathologic changes in the tissue. Selected unstained slides were submitted for Luxol fast blue-Periodic acid Schiff stains (LFB-PAS) to highlight myelin and collagenous connective tissues. "  "thermal injury, denoted by birefringence loss, can be detected in H&E-stained sections, the resolution of subtle changes is significantly improved when sections are stained with dyes that enhance the tissues’ natural birefringence. For example, picrosirius red (PSR) enhances collagen’s birefringence and has been shown to enable detection of thermal injury in collagen and muscle. "
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Is anyone familiar with setting up the protocol for recording spontaneous firing from midbrain dopaminergic neurons? (Also, we use Clampex-6). I have read couple of old foundational paper so far and the protocol is not mentioned anywhere in the paper. I am basically looking for the stimulation protocol and how much current was injected and in what frequency etc. Also, if the recording was done in a whole cell or cell-attached mode is also not mentioned in the paper. I am new into electrophysiology and I am not sure if the answers are there between the lines which I am not able to catch. Any help or suggestion will be highly appreciated. Also, if anyone is aware of any paper where it is given please let me know.
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In the loose cell-attached configuration, you are simply monitoring the ongoing spontaneous activity of the cell with minimal disruption/disturbance of the cell.  In the whole-cell configuration, you are dialyzing the entire interior of the cell, so the components of your internal solution will affect the activity of the cell (e.g. Cs vs K based internal, low vs high Cl, calcium buffering, etc).  As for loose cell-attached vs a high resistance cell-attached recording, it is difficult to prevent the membrane from rupturing under the patch when doing a high resistance cell-attached recording.  Thus, once you get even a small amount of membrane rupture, you are no longer in cell-attached mode and your internal will start to influence the activity of the cell.  This problem is greatly minimized (almost eliminated entirely) with in the loose-cell attached version.  This is also why most people use aCSF or something simple like NaHepes in the pipette during these recordings.
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Is it true that Biocytin is better distributet through a neuron, if it fires AP's and why would that be?
If I am using intracellular solution which is blocking AP's in my neuron, I heard from colleagues that the biocytin would not be distributed that well. Why is that so? How can AP's help the distribution of a fluochrome? Because of opening and closing channels and moving proteins troughout the cell?
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Hi Sarah,
I don't think that the firing itself helps the diffusion of biocytin. However depolarizing the pipette does help to push out biocytin, and that might explain why it looks like that the diffusion improves during firing.
For instance, to juxtacellularly fill neurons you need to apply positive current to the pipette, adjusting the amplitude to make the cell fire.
I'm attaching a couple of papers about juxtacellular filling with biocytin. 
In the whole cell configuration the main factors are the electrode resistance, the series resistance and the concentration of biocytin. I find that excitatory cells are very easy to fill, but inhibitory interneurons are much harder, in particular their axon is not always easy to preserve and/or fill (probably because is very thin).
Maximiliano
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There are many articles on the benefits of Imagery for poor readers'reading comprehension. Does anybody have any text, article, dissertation, talking about the counterpoint I mentioned in my question?
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Sidnei, maybe this paper can help you. All the reflections of Evelyn Arizpe about illustrations and picture books are very interesting.
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Under a blue light stimulation on ChR2-expressed interneurons, which is not decided whether through a pre- or post- synaptic mechanism modulating the neurotransmission of a disynaptic transmission close to it, the first phase of the EPSC was reduced, and PPR was found increased compared to without the light stimulation on those interneurons. 
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Essentially paired-pulse stimulus measures the short term plasticity characteristics of neurons. It is a characteristic of the probability of release of the presynaptic neuron and the type of receptors present at the postsynaptic neuron. PPR can be in both directions (facilitation or depression) and there are various explanations for what that means (as Bryan Krause mentioned).
A nice paper that goes over mechanisms for short term plasticity is: Fioravante & Regehr Current Opinion in Neurobiology 2011, 21:269–274
The postsynaptic receptor population should not have time to change during a paired pulse stimulation and if the treatment causes an effect by binding to the post-synaptic receptors, then the PPR should remain stable. While the amplitude of individual currents may be inhibited or enhanced by the treatment, the release of NT should be stable during the pulse and NT interactions with receptors should be equally effected by the postsynaptic change - thus the PPR is unchanged.
However, if the drug was interacting at the presynaptic site and changing the way the presynaptic neuron acts when it receives an action potential - its probability of release - then the way the presynaptic terminal responds to a paired pulse stimulus should also change - and the receptors see something different from before  - and PPR changes.
Its not conclusive, but a change in PPR after a drug treatment, indicates that the drug may be acting at the presynaptic side of the terminal.
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In patch clamp When I apply TTX + drug and I see a depolarization, it is a postsynaptic mechanism? But how can I check for the presynaptic mechanism?
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Not a lot of details to go on here.  Assuming you want to identify the mechanism underlying the depolarization, TTX application makes it less likely the drug is having a presynaptic effect (such as through increased spiking).  It also makes sense to analyze what the depolarization looks like.  If not smooth, then maybe it depends on presynaptic neurotransmitter release.  You could look at mini-EPSC frequency to verify drug is not having a presynaptic effect.  It would also make sense to block neurotransmitter receptors while you apply the drug (against GABA, AMPA, NMDA, mGluR), and if the depolarization stays intact, it is probably postsynaptic.
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We recently had a discussion if cell culturing lead to a higher basal activity compared to in vivo neurons which might occlude an effect in increased higher amplitude of the mEPSCs. You guys ever heard about this theory? It sounds reasonable to me but I can't find basic literature about it.
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Even more confusing is that during cell culture conditions, is it possible to have "autaptic" connection and to generate autaptic currents under "whole-cell" current recordings? I think that more importantly, because neuronal cell is rather small, how to determine cell-attached or whole-cell recordings is potentially important.
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I’m looking at the active zone for docking/priming/fusion between the nociceptor’s membrane and vesicles.
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Hi Laura!
Definitely, SNARE is involve in the nociceptive pathway. In our lab, we publish several papers speaking about its implication and even develop a peptide called DD04107 that interfere with the induced exocitosis of TRPV1.
Here you have some of this references:
An inhibitor of neuronal exocytosis (DD04107) displays long-lasting in vivo activity against chronic inflammatory and neuropathic pain
Bradykinin Induces TRPV1 Exocytotic Recruitment in Peptidergic Nociceptors
αCGRP is essential for algesic exocytotic mobilization of TRPV1 channels in peptidergic nociceptors
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I want to measure brain damage in EAE mice but i dont hve any experience about that
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I can send you LFB protocols on paraffin or frosen sections, as you prefere. It worked in rat and cat brain. But I am really curious how you will quantify the brain damage. And why don't you try Myelin Basic Protein Staining? LFB stains not only myelin. MBP is more specific
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By ma Na+ measurements.  I test the effect of  1mM D-Aspartat on my astrocytes they do not respond. I try with 8mM K+ and they show a drop down in the ratio. But later on also no effect on Aspartat? What is the reason for that. Actualy I should see something maybe to low concentrations. The Dye is fine i calibrated it propperly. Some guesses would be nice. If you need more Informations tell me
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In which case do you mean. WIth my Normal Ringer Perfusion or with the DY loding Hepes buffer oder With the K+ / D-Aspartat solution that i am using ???????
Thanks for you Answer
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Does anyone have experience in purifying plasma membrane from dorsal root ganglion? And even better, neuronal plasma membrane? We want to quantitate receptor membrane insertion within the ganglionic neurons.
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Hi Sean,
Well, I'm not a specialist in DRG purification, I only work on full mouse brain to isolate PM and Microsomal fractions,but you can use a subcellular fractionation protocol (multiple centrifugation). It works very well for me.
However, there is this article that describe a good method for DRG neurons.
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    We are doing experiments about rhythm in drosophila, and in data analysis part, we saw nearly all published papers use the software called FaasX, but we can't find it through the internet, so I'm wondering if someone have used it before, or is there anyone who can give me a clue, it would help a lot! I really appreciate for your help.
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We use Matlab with an specific toolbox, but if you want to use the FaasX I think you can download it from here:
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We work with Transgenic mice for Alzheimer's disease.
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I dislike the fluoroshield with DAPI. The DAPI remains in the tissue and causes many problems such as increase in background fluorescence. Rather, just buy a good mounting media for fluorescence without DAPI or make your own using Mowoil.
The DAPI stain can be done at the last step as follows: 1 min incubation in 1:1000 dilution  in PBS from 1 mg/mL stock sol in water and then wash thoroughly. DAPI is a very good fluorophore if used correctly.
Regards