From connelly.bill from gmail.com Mon Dec 1 01:22:45 2008 From: connelly.bill from gmail.com (Bill) Date: Mon Dec 1 12:08:36 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> Message-ID: <3051549b-d298-4d98-8499-6f57bec3b1c0@z28g2000prd.googlegroups.com> Mmm, there is something funny about CA3 cells, they've got to be one of the most delicate set of cells. The reason I reccomend the ascorbate is because of its important role in preventing the edema that occurs in slices. Check out: Ascorbate inhibits edema in brain slices. Brahma B, Forman RE, Stewart EE, Nicholson C, Rice ME. J Neurochem. 2000 Mar;74(3):1263-70 I've really noticed an improvement in the patchability of small cells if the slices are constantly incubated in ascorbate. I wonder about 3mM myo-inositol for similar reasons, though I've never tried that. Ah well, let me know how things go, I'm curious about the glycerol, I saw that paper, and it seemed a bit... amazing. On Dec 1, 4:32?am, Jose Guzman wrote: > Well, I agree with you, P21-30 are juvenile animals. I want to get > recordings of CA3 pyramidal neurons of the hippocampus. So far, I have > not much problems untill P18. However, due to the nature of my experiments, > I need to study older animals, and to have relative big number of cells > alive (only one or two cells per slice is not enough). Due to the extremely > high interconnectivity of those neurons, and the high NMDAR expression > I was thinking to use APV to avoid neurotoxicity due to Glu, but if > you say it's a waste of time I will think first about other solutions. > > I am carefully following Bischofberger et al, 2006. My cutting angle > seems to be OK, because after filling the cells with biocytine I can > clearly see both axon and dendrites (in animals < P18). My vibratome > is a Leica VT1200, so the state-of-the art. > > And YES, my cells look very crinkly (I was looking for that word!) > > I use a cutting solution in which NaCl is only partly replaced by > sucrosse (both for storage and cutting). No Glu-antagonists or > anti-oxidant agents like pyruvate or ascorbic acid. > > I will try soon the Glycerol-based solution, and additionally the > solution containing pyruvate and ascorbic acid (with my normal sucrosse > solution), and I will let you know how it was. > > Thank you very much in advance for your wisdom advices! > > Jose. > > On 2008-11-30, Bill wrote: > > > Hi Jose, > > > Just off the bat, lets remember that 21-30 days in not 'old'. Indeed, > > it is barely mature. Old is 18 month old rats. > > > Secondly, for 21-30 do not 'need' specialized cutting solution. The > > biggest change you see between slices from neonatal and ~30 day old > > brain slices is nothing to do there being more dead cells, it is > > simply that it is harder to see the neurons (though there are some > > areas of exception, e.g. highly mylinated areas). > > > The fact that the sucrose based solution didn't make a big difference > > informs you that depolarization toxicity is not your problem. > > > It would have been nice to know where your slices are from, so I could > > be certain, but assuming your working from some kind of cortical > > region: > > Sucrose is good. You might as well do it all the time >P14. AP5 is a > > waste of time, acute depolarization induced neurotoxicity is NMDA > > receptor independent. > > Incubate your solution at 35 degrees for half an hour after slicing, > > esp good for small cells. > > If your neurons are looking a bit desicated/crinkly/hard try 1mM > > Pyruvate and 3mM ascorbate in your incubating/holding aCSF > > What brand of slicer are you using? As your brain gets older, this > > gets far more important. You can cut nice slices from 12 day old brain > > with your hand and a razor blade, those neuros don't care. Make sure > > you're vibrotome has been serviced some time in the not too distant > > past. I've seen old Camden slicers that bounced around on the table > > because they needed salt crust chipped out of the motor and some > > lubrication. > > > But I stress this. If you're making slices from 21 day old animals, > > and your neurons look unhealthy (vs just harder to see) you are doing > > something wrong, indepedent of cutting solution. I worked on 8 week > > old mice for 2 years, and you can't get the neurons looking as round > > and full as 2 week old slices, and you can't get the neurons as > > visible (talk to someone who works on young cat thalamus if you want > > to hear about invisible neurons). But from a 21 day old animal, the > > neurons should still be like lovely round grapes, just harder to see > > than in a 14 day old. And if you're working with pyramidal cells, > > remember to keep your slice plane in the plane of the dendrites, those > > guys really hate having their dendrites cut. > > > Also, if you're working from an ultra mylinated structure, like the > > brain stem, it is vaguely impossible to get reasonable slices from 99% > > of that area in animals over ~21 days old, give or take a couple days. > > > I hope you haven't found this too didactic, but this is my experience > > (I'll probably get told this is completely wrong by someone else). > > > Oh finally, you can try intracardiac perfussion of your > > neuroprotective cutting solution. Its a pain, and its fiddley, I don't > > think it helps with the neuroprotection, but I think it helps makes > > the neurons in the slices more visible. > > > On Nov 28, 5:11?am, Jose Guzman wrote: > >> Hi everybody, > > >> I am currently working in hippocampal brain slices to do patch-clamp > >> electrophysiology. Due to some experimental issues, I will have to > >> record to neurons in old animals (P21-30). The problem is that the > >> neurons I need are very susceptible to hypoxia , so the preparation > >> requires some special cutting solutions. > > >> I collected the 3 major types of cutting solutions, and I would really > >> like to know if any of you tried these in his/her experiments . These > >> solutions have some minor variations in other publications (may contain > >> additionally APV or kyneuric acid to prevent glutamate excitotoxicity), > >> but basically are described as follows: > > >> ?1.- Sucrose based solution: total or partial replacement of NaCl > >> by Sucrose. (Aghajanian and Rasmussen, 1989, Moyer and Brown,1998). > > >> ?2.- Glycerol based solution: the same as the previous but with > >> ?Glycerol replacement. (Ye et al., 2006). > > >> ?3.- KGluconate based solution: without Ca2+ or Na+: > >> ?specially indicated for patching dendrites (Dugu? et al., 2005). > > >> ? ? ? ? ? ? A major problem I found is that these papers are not very > >> ? ? ? ? ? ? much cited in other publications (for example, Ye et al., > >> ? ? ? ? ? ? 2006 has only 6 citations). Personally, I found that > >> ? ? ? ? ? ? Sucrose-based solutions made an improvement (although not > >> ? ? ? ? ? ? very big) but I would really like to hear from your > >> ? ? ? ? ? ? experiences. > > >> ? ? ? ? ? ? Thank you very much for your feedback! From nin from neurohost.org Mon Dec 1 05:45:55 2008 From: nin from neurohost.org (Jose Guzman) Date: Mon Dec 1 12:08:41 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> <3051549b-d298-4d98-8499-6f57bec3b1c0@z28g2000prd.googlegroups.com> Message-ID: Thank you for the tips, I will try 3mM ascorbate late this week and possibly glycerol. I will let you know in every detail. And yes, the reason why not so many researchers are working on CA3 in slices is due to the difficulty of getting nice cells. I am preparing a document to describe the preparation of CA3 neurons in every little detail, and I will send you when it's finished. Thank you very much! Jose On 2008-12-01, Bill wrote: > Mmm, there is something funny about CA3 cells, they've got to be one > of the most delicate set of cells. > > The reason I reccomend the ascorbate is because of its important role > in preventing the edema that occurs in slices. Check out: > > Ascorbate inhibits edema in brain slices. > Brahma B, Forman RE, Stewart EE, Nicholson C, Rice ME. > J Neurochem. 2000 Mar;74(3):1263-70 > > I've really noticed an improvement in the patchability of small cells > if the slices are constantly incubated in ascorbate. > > I wonder about 3mM myo-inositol for similar reasons, though I've never > tried that. > > Ah well, let me know how things go, I'm curious about the glycerol, I > saw that paper, and it seemed a bit... amazing. > From luca_glb from hotmail.com Wed Dec 3 10:34:19 2008 From: luca_glb from hotmail.com (Luca Giliberto) Date: Wed Dec 3 12:52:20 2008 Subject: [Neuroscience] Actinomycin D Message-ID: Hi everyone. Question on Actinomycin D et similia. Does the degree of transcription inhibition of these compounds depend on the "copy number" of the gene/cDNA to be inhibited? E.g., if I transfect a cDNA-plasmid, with let's say a CMV type promoter, and go look mRNA from this cDNA and from an endogenous gene, should I expect same the amount of mRNA (low), or the transfected gene, being "more", has more chances to escape the inhibition and get transcribed? Thanks. Luca. From nin from neurohost.org Thu Dec 4 06:17:41 2008 From: nin from neurohost.org (Jose Guzman) Date: Thu Dec 4 12:30:51 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> <3051549b-d298-4d98-8499-6f57bec3b1c0@z28g2000prd.googlegroups.com> Message-ID: As you pointed out, when I used a mixed NaCl/Sucrosse solution with ascorbic acid (3mM), the quality of the slices were slightly better, but the cells were much more easy to patch. I am currently thinking to put ascorbic acid during my recording solution (without Sucrosse). On the other hand, I used pyruvic acid (1mM). Is there any reasons other than "neuroprotection" involved in the effect of pyruvic acid on brain slice quality? I did not find any conclussive report. Next week I will do the same cutting solution but with glycerol (mixed NaCl/Glycerol + Ascorbic acid + pyruvate). I will let you know as soon as possible. Best wishes and thank you very much! Jose. On 2008-12-01, Bill wrote: > Mmm, there is something funny about CA3 cells, they've got to be one > of the most delicate set of cells. > > The reason I reccomend the ascorbate is because of its important role > in preventing the edema that occurs in slices. Check out: > > Ascorbate inhibits edema in brain slices. > Brahma B, Forman RE, Stewart EE, Nicholson C, Rice ME. > J Neurochem. 2000 Mar;74(3):1263-70 > > I've really noticed an improvement in the patchability of small cells > if the slices are constantly incubated in ascorbate. > > I wonder about 3mM myo-inositol for similar reasons, though I've never > tried that. > > Ah well, let me know how things go, I'm curious about the glycerol, I > saw that paper, and it seemed a bit... amazing. > > On Dec 1, 4:32?am, Jose Guzman wrote: >> Well, I agree with you, P21-30 are juvenile animals. I want to get >> recordings of CA3 pyramidal neurons of the hippocampus. So far, I have >> not much problems untill P18. However, due to the nature of my experiments, >> I need to study older animals, and to have relative big number of cells >> alive (only one or two cells per slice is not enough). Due to the extremely >> high interconnectivity of those neurons, and the high NMDAR expression >> I was thinking to use APV to avoid neurotoxicity due to Glu, but if >> you say it's a waste of time I will think first about other solutions. >> >> I am carefully following Bischofberger et al, 2006. My cutting angle >> seems to be OK, because after filling the cells with biocytine I can >> clearly see both axon and dendrites (in animals < P18). My vibratome >> is a Leica VT1200, so the state-of-the art. >> >> And YES, my cells look very crinkly (I was looking for that word!) >> >> I use a cutting solution in which NaCl is only partly replaced by >> sucrosse (both for storage and cutting). No Glu-antagonists or >> anti-oxidant agents like pyruvate or ascorbic acid. >> >> I will try soon the Glycerol-based solution, and additionally the >> solution containing pyruvate and ascorbic acid (with my normal sucrosse >> solution), and I will let you know how it was. >> >> Thank you very much in advance for your wisdom advices! >> >> Jose. >> >> On 2008-11-30, Bill wrote: >> >> > Hi Jose, >> >> > Just off the bat, lets remember that 21-30 days in not 'old'. Indeed, >> > it is barely mature. Old is 18 month old rats. >> >> > Secondly, for 21-30 do not 'need' specialized cutting solution. The >> > biggest change you see between slices from neonatal and ~30 day old >> > brain slices is nothing to do there being more dead cells, it is >> > simply that it is harder to see the neurons (though there are some >> > areas of exception, e.g. highly mylinated areas). >> >> > The fact that the sucrose based solution didn't make a big difference >> > informs you that depolarization toxicity is not your problem. >> >> > It would have been nice to know where your slices are from, so I could >> > be certain, but assuming your working from some kind of cortical >> > region: >> > Sucrose is good. You might as well do it all the time >P14. AP5 is a >> > waste of time, acute depolarization induced neurotoxicity is NMDA >> > receptor independent. >> > Incubate your solution at 35 degrees for half an hour after slicing, >> > esp good for small cells. >> > If your neurons are looking a bit desicated/crinkly/hard try 1mM >> > Pyruvate and 3mM ascorbate in your incubating/holding aCSF >> > What brand of slicer are you using? As your brain gets older, this >> > gets far more important. You can cut nice slices from 12 day old brain >> > with your hand and a razor blade, those neuros don't care. Make sure >> > you're vibrotome has been serviced some time in the not too distant >> > past. I've seen old Camden slicers that bounced around on the table >> > because they needed salt crust chipped out of the motor and some >> > lubrication. >> >> > But I stress this. If you're making slices from 21 day old animals, >> > and your neurons look unhealthy (vs just harder to see) you are doing >> > something wrong, indepedent of cutting solution. I worked on 8 week >> > old mice for 2 years, and you can't get the neurons looking as round >> > and full as 2 week old slices, and you can't get the neurons as >> > visible (talk to someone who works on young cat thalamus if you want >> > to hear about invisible neurons). But from a 21 day old animal, the >> > neurons should still be like lovely round grapes, just harder to see >> > than in a 14 day old. And if you're working with pyramidal cells, >> > remember to keep your slice plane in the plane of the dendrites, those >> > guys really hate having their dendrites cut. >> >> > Also, if you're working from an ultra mylinated structure, like the >> > brain stem, it is vaguely impossible to get reasonable slices from 99% >> > of that area in animals over ~21 days old, give or take a couple days. >> >> > I hope you haven't found this too didactic, but this is my experience >> > (I'll probably get told this is completely wrong by someone else). >> >> > Oh finally, you can try intracardiac perfussion of your >> > neuroprotective cutting solution. Its a pain, and its fiddley, I don't >> > think it helps with the neuroprotection, but I think it helps makes >> > the neurons in the slices more visible. >> >> > On Nov 28, 5:11?am, Jose Guzman wrote: >> >> Hi everybody, >> >> >> I am currently working in hippocampal brain slices to do patch-clamp >> >> electrophysiology. Due to some experimental issues, I will have to >> >> record to neurons in old animals (P21-30). The problem is that the >> >> neurons I need are very susceptible to hypoxia , so the preparation >> >> requires some special cutting solutions. >> >> >> I collected the 3 major types of cutting solutions, and I would really >> >> like to know if any of you tried these in his/her experiments . These >> >> solutions have some minor variations in other publications (may contain >> >> additionally APV or kyneuric acid to prevent glutamate excitotoxicity), >> >> but basically are described as follows: >> >> >> ?1.- Sucrose based solution: total or partial replacement of NaCl >> >> by Sucrose. (Aghajanian and Rasmussen, 1989, Moyer and Brown,1998). >> >> >> ?2.- Glycerol based solution: the same as the previous but with >> >> ?Glycerol replacement. (Ye et al., 2006). >> >> >> ?3.- KGluconate based solution: without Ca2+ or Na+: >> >> ?specially indicated for patching dendrites (Dugu? et al., 2005). >> >> >> ? ? ? ? ? ? A major problem I found is that these papers are not very >> >> ? ? ? ? ? ? much cited in other publications (for example, Ye et al., >> >> ? ? ? ? ? ? 2006 has only 6 citations). Personally, I found that >> >> ? ? ? ? ? ? Sucrose-based solutions made an improvement (although not >> >> ? ? ? ? ? ? very big) but I would really like to hear from your >> >> ? ? ? ? ? ? experiences. >> >> >> ? ? ? ? ? ? Thank you very much for your feedback! > From usenet02 from out-of-phase.de Thu Dec 4 13:36:30 2008 From: usenet02 from out-of-phase.de (Christian Wilms) Date: Thu Dec 4 14:05:50 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> Message-ID: <1irfstn.izm2vh1fpireaN%usenet02@out-of-phase.de> Hey Jose! I thought you were foraging in vivo now :) > I use a cutting solution in which NaCl is only partly replaced by > sucrosse (both for storage and cutting). No Glu-antagonists or > anti-oxidant agents like pyruvate or ascorbic acid. I've never used different solutions for slicing and recording, as cerebellum in juveline animals is quite straight forward. I know Bernardo Sabatini's group tends to use choline-chloride based solutions for slicing. What I've seen people in our lab do (if they needed perfect preservation of fine structures) is to store the slices in the sucrose solution after slicing and subsequently _slowly_ (takes around 20 minutes) replace the solution with regular ACSF. Good luck with your experiments, Christian From kongzd from gmail.com Thu Dec 4 14:51:15 2008 From: kongzd from gmail.com (kongzd@gmail.com) Date: Thu Dec 4 17:15:41 2008 Subject: [Neuroscience] which dye do I choose? It shouldnot cross the synapse , and keep the fluorescence for a long time (a few months)? Message-ID: <4eeefbbf-ec59-4b5a-8876-de9ce99caedc@c36g2000prc.googlegroups.com> Hi my experiment is transplanted the neurons to the nerve tissue of host. Now I want to observe the synapse between the transplanted neurons and host neurons. So before transplantation, i go to find a dye to staining the transplanted neurons. this dye shouldnot cross the synapse,so I can obsevre the synapse in electron scope. and the experimentsin vivo lasts a long time ,I want to it keep the fluorescence for about 4 month. is there a dye recommended? does DiI or Dio meet the requirement? I really donnot konw if DiI can cross the synapse. thank you. From biocjh from gmail.com Fri Dec 5 03:41:31 2008 From: biocjh from gmail.com (jh) Date: Fri Dec 5 11:58:03 2008 Subject: [Neuroscience] help on neuron-virus interaction Message-ID: Dear all? I am writing to seek for help on neurovirology. Anyone know leading virologists whose interest is focus on the neuron-virus interaction. rgds! jianhao From wwt0657 from gmail.com Sun Dec 7 08:55:33 2008 From: wwt0657 from gmail.com (wwt) Date: Sun Dec 7 13:10:13 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> <1irfstn.izm2vh1fpireaN%usenet02@out-of-phase.de> Message-ID: <40ad1699-0572-415e-8476-64bb59efb406@c36g2000prc.googlegroups.com> I read a book introduced the role of ascorbate. But in the book, the author used two words: sodium ascorbate and ascorbate acid. Do you know which one work on acsf for inhibting edema? I had cutted inferior olive slice in 14d-22d rats. The health of slice of >18d is too bad. All of cells are swelled. The cells of 14d are good for recording. I had asked Yosi Yarom's Research Lab for the methods of brain stem. They suggest intracardiac perfussion is the first step. After perfussion, they use a strange solution for cutting slice. The solution is (mM):glucose 10, KCl 5, NaHCO3 26, CaCl2 2.4, KH2PO4 1.2, MgSO4 1.3, Surcose 124. The osmotic pressure is about 200. After cutting, they use normal acsf replace the cutting solution. I had asked them for the reason of low osmotic cutting solution. They also did know the reason. But they think it work. Do you know heard about it and know the reason? I tried the method according their advises. But the slices are still unhealth. I think there are other tips that I don't notice, specially in taking out the brain stem. From claudioez from gmail.com Mon Dec 8 07:13:36 2008 From: claudioez from gmail.com (=?ISO-8859-1?Q?Claudio_Elgueta_Zu=F1iga?=) Date: Mon Dec 8 12:19:14 2008 Subject: [Neuroscience] Looking for a microscope camera Message-ID: Hey guys: I need to buy a camera to take some pictures of my Lucifer Yellow filled cells after myvoltage clamp experiments, using a Olympus microscope with infrared DIC optic. Anyone knows about a cheap digital camera with good infrared sensitivity and a decent resolution?. Currently I?m using a analog security camera, but it has a poor resolution and the software I have to acquire images is not so good also. Best regards, Claudio. -- Claudio Elgueta Bioqu?mico Universidad de Chile Programa doctorado en ciencias menci?n neurociencias, Universidad de Valpara?so. Centro de neurociencias de Valpara?so. Gran Breta?a 1111 032-2508186 From usenet02 from out-of-phase.de Mon Dec 8 09:12:37 2008 From: usenet02 from out-of-phase.de (Christian Wilms) Date: Mon Dec 8 12:19:22 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> <1irfstn.izm2vh1fpireaN%usenet02@out-of-phase.de> <40ad1699-0572-415e-8476-64bb59efb406@c36g2000prc.googlegroups.com> Message-ID: <1irluks.19tkhdp181fzuoN%usenet02@out-of-phase.de> Especially when switching solutions from hypoosmol to "normal ACSF" it is very important to slowly exchange solution. Ideally, a peristaltic pump will be pumping regular ACSF into the bath containing the finished slices while remove excess solution at the same time. A good rate should be 2 to 5 mL per minute. If you use ascorbic acid or its salt form sodium ascorbate shouldn't matter, as long as you adjust the pH properly. In both cases the two forms will be present in an equilibrium that will depend on the pH of your solution (according to the Henderson-Hasselbalch equation). h2h, Christian From wwt0657 from gmail.com Mon Dec 8 20:14:42 2008 From: wwt0657 from gmail.com (wwt) Date: Mon Dec 8 22:12:17 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> <1irfstn.izm2vh1fpireaN%usenet02@out-of-phase.de> <40ad1699-0572-415e-8476-64bb59efb406@c36g2000prc.googlegroups.com> <1irluks.19tkhdp181fzuoN%usenet02@out-of-phase.de> Message-ID: <692343e8-c230-4d80-93a9-c840b3d622d1@g17g2000prg.googlegroups.com> On Dec 8, 10:12?pm, usene...@out-of-phase.de (Christian Wilms) wrote: > Especially when switching solutions from hypoosmol to "normal ACSF" it > is very important to slowly exchange solution. Ideally, a peristaltic > pump will be pumping regular ACSF into the bath containing the finished > slices while remove excess solution at the same time. A good rate should > be 2 to 5 mL per minute. > > If you use ascorbic acid or its salt form sodium ascorbate shouldn't > matter, as long as you adjust the pH properly. In both cases the two > forms will be present in an equilibrium that will depend on the pH of > your solution (according to the Henderson-Hasselbalch equation). > > h2h, Christian Thanks for your answer. But I still don't understand the reason for using hypoosmol solution. What good do hypoosmol solution? From usenet02 from out-of-phase.de Tue Dec 9 06:17:31 2008 From: usenet02 from out-of-phase.de (Christian Wilms) Date: Tue Dec 9 08:36:30 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> <1irfstn.izm2vh1fpireaN%usenet02@out-of-phase.de> <40ad1699-0572-415e-8476-64bb59efb406@c36g2000prc.googlegroups.com> <1irluks.19tkhdp181fzuoN%usenet02@out-of-phase.de> <692343e8-c230-4d80-93a9-c840b3d622d1@g17g2000prg.googlegroups.com> Message-ID: <1irohwr.9l8b4xnpx33eN%usenet02@out-of-phase.de> wwt wrote: > Thanks for your answer. But I still don't understand the reason for > using hypoosmol solution. What good do hypoosmol solution? Sorry, I gotta pass on that as well. In the end I guess it's a question of what works for you. Slicing is unfortunatelt full of heuristics. :) Best, Christian From nin from neurohost.org Tue Dec 9 12:31:27 2008 From: nin from neurohost.org (Jose Guzman) Date: Tue Dec 9 17:26:30 2008 Subject: [Neuroscience] Re: Cuttting solutions for electrophysiology References: <5a7ed4e7-77ab-4e29-bea9-b6c7d4d19152@k36g2000pri.googlegroups.com> <1irfstn.izm2vh1fpireaN%usenet02@out-of-phase.de> <40ad1699-0572-415e-8476-64bb59efb406@c36g2000prc.googlegroups.com> <1irluks.19tkhdp181fzuoN%usenet02@out-of-phase.de> <692343e8-c230-4d80-93a9-c840b3d622d1@g17g2000prg.googlegroups.com> Message-ID: Usually intracellular solution has higher osmolarity. If the osmolarity of your pipette solution is a little bit higher than the osmolarity of your bath solution, (say 310 versus 300 mOms) cells will swell slightly and they are easier to path. Regards! Jose. > Thanks for your answer. But I still don't understand the reason for > using hypoosmol solution. What good do hypoosmol solution? From rajasekaran.karthik from gmail.com Tue Dec 9 14:36:04 2008 From: rajasekaran.karthik from gmail.com (Karthik) Date: Tue Dec 9 17:26:36 2008 Subject: [Neuroscience] does spermine impede patch clamping neurons? Message-ID: Hi, I am trying to record rectification of AMPA currents, and am having problems with the quality of patch clamping if I add 100 microM of spermine to my pipette solution. I tried by first back filling my pipette with the regular internal and then filling with spermine containing internal, in which case the quality of seal degrades within a few minutes of patching the cell. Would appreciate any input on this problem. Thanks a lot! Karthik From kirsch from bccn.uni-freiburg.de Mon Dec 15 09:52:37 2008 From: kirsch from bccn.uni-freiburg.de (Janina Kirsch) Date: Mon Dec 15 14:50:58 2008 Subject: [Neuroscience] PhD-Positions in Neurotechnology in Freiburg, Germany Message-ID: <850DD0102DB94982BFFD0FDAC71DB6CB@janina> 11 PhD-Positions in Neurotechnology Logo_BFNT_web.jpg Neurotechnology is a rapidly growing field of research, thriving to repair, replace or support physical functions affected by nervous system diseases by technical means. At the Bernstein Focus for Neurotechnology (BFNT) we exploit electrical and chemical signals recorded in the brain to control external or implanted devices. The BFNT efforts are organized in projects on fundamental neurotechnological research, the development of new techniques, clinical testing and neuroethical considerations. PhD-Positions are available in the following projects: . Voluntary control of neuronal correlations for BMIs (Project A2) . Co-Adaptivity of brain and computer: Interaction of two learning signals (Project A3) . Adaptive interception of epileptic events based on predictive network dynamics (Project A4) . Development, prototyping, fabrication and testing of individualized and long-term safe electrode arrays (Project B1) . Multielectrode arrays with adjustable stiffness (Project B2) . Improving microelectrodes for single cell recordings in the human brain (Project B3) . NETCAD - An environment for 3D structural modeling of biological neuronal networks (Project B5) . Real time metabolite sensing for feedback control of behaviour in neurological disorders (Project C4) . Multicontact stimulation techniques for interventions in human focal epilepsy (Project C5) Further details on: www.bfnt.uni-freiburg.de/jobs Contact: -- Dr. Janina Kirsch -- Coordinator for the BCCN Teaching & Training Programs Bernstein Center for Computational Neuroscience Albert-Ludwigs-University Freiburg Hansastr. 9a D - 79104 Freiburg Germany Phone: +49 (0) 761 203-9575 Fax: +49 (0) 761 203-9559 Email: kirsch @bcf.uni-freiburg.de Web: www.bcf.uni-freiburg.de From kirsch from bccn.uni-freiburg.de Mon Dec 15 09:52:37 2008 From: kirsch from bccn.uni-freiburg.de (Janina Kirsch) Date: Mon Dec 15 14:51:10 2008 Subject: [Neuroscience] Postdoc-Positions in Neurotechnology in Freiburg, Germany Message-ID: <8F26EB5E3DAB427892AD1F1C97A05CFD@janina> 5 Postdoc-Positions in Neurotechnology Logo_BFNT_web.jpg Neurotechnology is a rapidly growing field of research, thriving to repair, replace or support physical functions affected by nervous system diseases by technical means. At the Bernstein Focus for Neurotechnology (BFNT) we exploit electrical and chemical signals recorded in the brain to control external or implanted devices. The BFNT efforts are organized in projects on fundamental neurotechnological research, the development of new techniques, clinical testing and neuroethical considerations. Postdoc-Positions are available in the following projects: . Co-Adaptivity of brain and computer: Interaction of two learning signals (Project A3) . Defined stimulus-response functions in active networks (Project A5) . Improving microelectrodes for single cell recordings in the human brain (Project B3) . NEURONLINE - System for the on-site/on-line monitoring of neuronal population activity (Project B4) . Neuroethics & Neurotechnology: Emerging questions from hybrid brains (Project C6) Further details on: www.bfnt.uni-freiburg.de/jobs Contact: -- Dr. Janina Kirsch -- Coordinator for the BCCN Teaching & Training Programs Bernstein Center for Computational Neuroscience Albert-Ludwigs-University Freiburg Hansastr. 9a D - 79104 Freiburg Germany Phone: +49 (0) 761 203-9575 Fax: +49 (0) 761 203-9559 Email: kirsch @bcf.uni-freiburg.de Web: www.bcf.uni-freiburg.de From connelly.bill from gmail.com Sat Dec 20 21:44:58 2008 From: connelly.bill from gmail.com (Bill) Date: Sun Dec 21 14:31:11 2008 Subject: [Neuroscience] Re: does spermine impede patch clamping neurons? References: Message-ID: This wont be much help Karthik, but no, I've patched with spermine in my pipette for 3 weeks, and I didn't find it effected initial seal quality, or longevity of recordings. On Dec 10, 8:36?am, Karthik wrote: > Hi, I am trying to record rectification of AMPA currents, and am > having problems with the quality of patch clamping if I add 100 microM > of spermine to my pipette solution. I tried by first back filling my > pipette with the regular internal and then filling with spermine > containing internal, in which case the quality of seal degrades within > a few minutes of patching the cell. Would appreciate any input on this > problem. Thanks a lot! > Karthik From connelly.bill from gmail.com Sat Dec 20 21:52:21 2008 From: connelly.bill from gmail.com (Bill) Date: Sun Dec 21 14:31:17 2008 Subject: [Neuroscience] Re: Looking for a microscope camera References: Message-ID: <167a8560-bd87-4e8e-9513-4111229d3bae@o4g2000pra.googlegroups.com> Hi Claudio, It's nice to hear I'm not the only one who uses a security camera for IR-DIC! If you just want to take pictures, then the DAGE-IR100 is excellent. Its great for IR-DIC, and has excellent sensitivity for its price. It doesn't come with any software/capture card etc, however now days you can get a capture card with a video port for ~$50. You can set up digital pulses to control exposure time, but you probably wont need to, to capture lucifer yellow. If you're looking for a camera built for this kind of stuff, and it will allow you do to ROI analysis, binning, etc for Ca2+ experiments, one of the cheapest cameras that I know of is the Diagnostic Instruments Pursuit range, which are about 11,000 US. If someone knows a better deal let everyone know. On Dec 9, 1:13?am, "Claudio Elgueta Zu?iga" wrote: > Hey guys: > > ? ? ? ? ? ? ? ? I need to buy a camera to take some pictures of my Lucifer > Yellow filled cells after myvoltage clamp experiments, using a Olympus > microscope with infrared DIC optic. Anyone knows about a cheap digital > camera with good infrared sensitivity and a decent resolution?. Currently > I?m using a analog security camera, but it has a poor resolution and the > software I have to acquire images is not so good also. > > Best regards, Claudio. > > -- > > Claudio Elgueta > > Bioqu?mico Universidad de Chile > > Programa doctorado en ciencias menci?n neurociencias, Universidad de > Valpara?so. > > Centro de neurociencias de Valpara?so. > > Gran Breta?a 1111 > > 032-2508186 From connelly.bill from gmail.com Wed Dec 24 00:57:10 2008 From: connelly.bill from gmail.com (Bill) Date: Wed Dec 24 13:33:04 2008 Subject: [Neuroscience] Re: which dye do I choose? It shouldnot cross the synapse , and keep the fluorescence for a long time (a few months)? References: <4eeefbbf-ec59-4b5a-8876-de9ce99caedc@c36g2000prc.googlegroups.com> Message-ID: On Dec 5, 8:51?am, kon...@gmail.com wrote: > Hi > my experiment is transplanted the neurons to the nerve tissue of host. > Now I want to observe the synapse between the transplanted neurons and > host neurons. So before transplantation, i go to find a dye to > staining the ?transplanted neurons. > this dye shouldnot cross the synapse,so I can obsevre the synapse in > electron scope. > and the experimentsin vivo lasts a long time ,I want to it keep the > fluorescence for about 4 month. > > is there a dye recommended? does DiI or Dio meet the requirement? I > really donnot konw if DiI can cross the synapse. > thank you. Sounds like you're going to need a transgenic animal to me. GFP under the NeuN promoter or something. Stick glowing neurons into the head of the wild-type animal. I'm no expert, but it strikes me that most dyes will be metabolized and cleared in live tissue over 4 months. From kldickson from wisc.edu Sat Dec 27 14:51:47 2008 From: kldickson from wisc.edu (KATHARINE LEAH DICKSON) Date: Sat Dec 27 15:02:23 2008 Subject: [Neuroscience] Neuroscience programs: Got a list, need tips. Message-ID: Hello! This is my first post. I have a list of some PhD programs I'd like to apply to, but I need a little bit more advice as to how to narrow it down. I'm an undergrad and have a little time before I start applying to grad schools, so I'm narrowing them down ahead of time before I start applying so I won't spend excess time worrying about where to apply when I start applying. There are apparently a whole LOT of places, more than I expected (around 30), concerning sufficiently related projects to what I want to do (although what 'sufficiently' is, I'm not completely sure, and I need a better idea of how to really refine that). How do I narrow it down to a reasonable number, and what is a reasonable number to apply to? I've heard anywhere from as few as 5 to as many as 15. Thanks - kldickson@wisc.edu From thome.alex from gmail.com Sun Dec 28 12:21:31 2008 From: thome.alex from gmail.com (Alex Thome) Date: Sun Dec 28 15:01:40 2008 Subject: [Neuroscience] Re: Neur-sci Digest, Vol 43, Issue 11 In-Reply-To: <200812281705.mBSH54f06469@net.bio.net> References: <200812281705.mBSH54f06469@net.bio.net> Message-ID: Hi Katharine, good question and a great place to ask it. I would say 5 is certainly good, and 15 is better given that during times of economic hardship people seek refuge in academia. As far as picking the best place goes, I would recommend contacting some of the PI's you are interested in and having an informal discussion with them regarding projects/positions. One good place to do this would be to go to the annual SFN meeting and try to meet as many of your chosen PI's (as well as their post-doc and grad-students) in person. Having a personal meeting, makes you more memorable from the stacks of 100's of applications they may have to sift through, but also will tell you whether there may be a personality conflict. Being a grad student that's about as much as I can tell you, feel free to contact me if you have any further questions. Cheers, Alex ________________________________ Graduate Student/Research Assistant McKnight Brain Institute & ARL Neural Systems Memory and Aging M: (520) 891-2936 http://embi.nsma.arizona.edu Not knowing is true knowledge. Presuming to know is a disease. First realize that you are sick. Then you can move towards health. - Lao Tze ---------------------------------------------------------------------- > > Message: 1 > Date: Sat, 27 Dec 2008 13:51:47 -0600 > From: KATHARINE LEAH DICKSON > Subject: [Neuroscience] Neuroscience programs: Got a list, need tips. > To: neur-sci@magpie.bio.indiana.edu > Message-ID: > Content-Type: text/plain; charset=us-ascii > > Hello! This is my first post. > > I have a list of some PhD programs I'd like to apply to, but I need a > little bit more advice as to how to narrow it down. I'm an undergrad and > have a little time before I start applying to grad schools, so I'm narrowing > them down ahead of time before I start applying so I won't spend excess time > worrying about where to apply when I start applying. > > There are apparently a whole LOT of places, more than I expected (around > 30), concerning sufficiently related projects to what I want to do (although > what 'sufficiently' is, I'm not completely sure, and I need a better idea of > how to really refine that). How do I narrow it down to a reasonable number, > and what is a reasonable number to apply to? I've heard anywhere from as > few as 5 to as many as 15. > > Thanks - > > kldickson@wisc.edu > > > > ------------------------------ > > _______________________________________________ > Neur-sci mailing list > Neur-sci@net.bio.net > http://www.bio.net/biomail/listinfo/neur-sci > > End of Neur-sci Digest, Vol 43, Issue 11 > **************************************** > From kldickson from wisc.edu Mon Dec 29 13:47:17 2008 From: kldickson from wisc.edu (KATHARINE LEAH DICKSON) Date: Mon Dec 29 13:55:01 2008 Subject: [Neuroscience] Reputation of programs Message-ID: Also, another question: I know the reputations of some programs on my list, but I don't know the reputations of others. Can you tell me about the neuroscience programs at UIUC, Dartmouth, UT-San Antonio, UC-Riverside, UC-Davis, Northwestern, UAlabama-Birmingham, and Vanderbilt? Katharine Leah Dickson kldickson@wisc.edu