P/Q-type calcium channels mediate the activity-dependent feedback of syntaxin-1A
Authors: H Guthrie
J E McRory
K G Sutton
T P Snutch
T H Murphy
Journal: Nature (1999)
Discipline: Multidisciplinary
Keywords: intracellular calcium concentrations; gene expression; neurons; L-type channels; transcription factor CREB
Affiliations: Biotechnology Laboratory, Dept Psychiatry University of British Columbia, Vancouver, Canada.

Abstract:  Spatial and temporal changes in intracellular calcium concentrations are critical for controlling gene expression in neurons. In many neurons, activity-dependent calcium influx through L-type channels stimulates transcription that depends on the transcription factor CREB by activating a calmodulin-dependent pathway. Here we show that selective influx of calcium through P/Q-type channels is responsible for activating expression of syntaxin-1A, a presynaptic protein that mediates vesicle docking, fusion and neurotransmitter release. The initial P/Q-type calcium signal is amplified by release of calcium from intracellular stores and acts through phosphorylation that is dependent on the calmodulin-dependent kinase CaM K II/IV, protein kinase A and mitogen-activated protein kinase kinase. Initiation of syntaxin-1A expression is rapid and short-lived, with syntaxin-1A ultimately interacting with the P/Q-type calcium channel to decrease channel availability. Our results define an activity-dependent feedback pathway that may regulate synaptic efficacy and function in the nervous system. 

Retracted: No

PubPeer comments:
 Figure 1B. * Blue and cyan boxes: The PBS and LGS-H panels appear to show clear regions of overlap, at both magnifications. This appears to be a key figure in the conclusions of the paper that Liang-Ge-San protects zebrafish against LPS-induced inflammation.  , 
 Dear Dr. Bik, I am Junshan Liu, the author of the article u201cLiang-Ge-San, a classic traditional Chinese medicine formula, attenuates acute inflammation in zebrafish and RAW 264.7 cellsu201d. Sorry for the late reply, since Dr. Zhouu2019s registration (zhoull2020@hust.edu.cn) is not verified by an admin up to now. She authorized me to reply for her. After receiving your comment, we carefully checked the experimental records of Figure 1B. Unfortunately, we found that we mistakenly used the pictures of PBS group as the LGS-H group when processing pictures. The corrected raw data is attached below. We sincerely apologize for this unintentional error and confirm that the mistake would not affect the conclusions of this study. At the same time, we have contacted the journal to correct the misused pictures. We greatly appreciate for your concern.  , 
 #2 Dear Dr. Junshan Liu, While you are looking into this publication, could I perhaps advice you to also look into the Kaplan-Meier plot in Figure 1A.  There is some quite remarkable behavior in this figure. I wonder if there might be a methodological explanation for it? Firstly, there is the PBS group where larvae die off exactly five at a time throughout the experiment. Now, obviously it is impossible to make observations in an experiment like this frequently enough to be able to see when each individual larvae dies. But it seems like a coincidence of astronomical proportions that they would have been recorded at multiples of five throughout the experiment. Could there be a methodological explanation? Moreover, in the LPS-LGS-L group, the larvae, again, die off in cohorts of identical size. Using an on-screen ruler, we can tell that the group is reduced in increments of 12.5 %. This is however a bit of a problem, since the group is listed at n = 50. The larvae seem to die off, 6.25 at a time, which is impossible. Please look into what may have gone wrong here. It would also be nice if you made the raw data available. , 
 Dear Dr. Bik, I am Hongling Zhou, the first author of article u201cLiang-Ge-San, a classic traditional Chinese medicine formula, attenuates acute inflammation in zebrafish and RAW 264.7 cellsu201d. Sorry for the late reply, I had just undergone a surgery in the past two days. After receiving your comment, we carefully checked the experimental records of Figure 1B. Unfortunately, we found that we mistakenly used the pictures of PBS group as the LGS-H group when processing pictures. The corrected raw data is attached below. We sincerely apologize for this unintentional error and confirm that the mistake would not affect the conclusions of this study. At the same time, we have contacted the journal to correct the misused pictures. We greatly appreciate for your concern.  , 
 Dear Dr. Malacotricha, I am Hongling Zhou, the first author of article u201cLiang-Ge-San, a classic traditional Chinese medicine formula, attenuates acute inflammation in zebrafish and RAW 264.7 cellsu201d. Thank you for your comments on the above article. After carefully evaluated your critical comments and thoughtful suggestions, we have the following responses: 1. Zebrafish in the PBS group did not die. The black survival curve indicated that the survival rate of PBS-injected zebrafish was 100% within 72 hours. 2. The survival condition of zebrafish in each group was observed and recorded at specific intervals, which was determined by pre-experiments. 3. Unfortunately, we found a typing mistake in the figure legends of Figure 1A, the group is listed at n=10, but not n=50. We are very sorry to this unintentional mistake. The corrected figure and raw data are attached as following. We sincerely apologize for this unintentional error and confirm that the mistake would not affect the conclusions of this study. The corresponding corrigendum had been sent to the editor of the journal. We would like to express our great appreciation to you for comments on our paper, which are valuable and helpful. And also alert us to strictly check all data and manuscript in the future submission.   , 
 #5 1. PBS group 100% survival -- agreed, I think #3 may have been a typo. 2.  ...recorded at specific intervals, which was determined by pre-experiments. How did you determine these intervals? The intervals seem to vary from 4h to 12h, and I don t see any regularity. 3. The plot you show there is fundamentally different from the one in the paper. The LPS+LGS-H group, for example went from about 80% survival to 90% survival. Also, you are now claiming that at each of the intervals you found, at most, one dead individual per colony, never two, never three, even when the intervals differed by a factor of 3, and even when one colony lost all of the individuals?? Really? This error is much more than a  typing mistake in the figure legends of Figure 1A, the group is listed at n=10, but not n=50 , the data is different!   , 
 #6 1. My apologies for the typo, I did of course mean the LPS group in my previous post. An sample size of 10 does indeed explain some of the behavior of the published figure. 2 3. I would like to second the confusion of the previous commenter, Atheropla Decaspila, however. It is rather remarkable that the haphazardly distributed observations should happen to fall in such a way that only *exactly* one embryo will have died in each group - never two or three. in the period between 31 and 35 hpi, four embryos died, but they all remarkably fell into different experimental groups. I wonder if there is anything in the methods that might make this all seem less ... serendipitous? Dear Drs Hongling Zhou and Junshan Liu, could you perhaps for the benefit of our understanding also show the raw data that produced the erroneous previous Kaplan-Meier plot? I am similarly confused as to what has changed and why. , 
 Dear Dr. Decaspila and Dr. Malacotricha, thank you for your comments on u201cLiang-Ge-San, a classic traditional Chinese medicine formula, attenuates acute inflammation in zebrafish and RAW264.7 cellsu201d. On behalf of my co-authors, I would like to give a detailed explanation below. 1. Either irregular or regular interval is acceptable in the observation of survival ratio. In our pre-experiment, we observed the survival of larvae in LPS group. Once one individual larva in the group was dead, the time point was recorded. Under 72 h continuous observation, we recorded death time of each larva. Then, in the following study, these time points (as shown in attached figure A, first column) were further used to observe the survival conditions of all groups. Therefore, the observation interval is irregular. 2. As described above, the time interval was determined according to the death of larvae in LPS group. Therefore, only one larva was dead at a time point in LPS group. Moreover, it is easy to understand that only one larva or none was dead in other groups due to dexamethasone and LGS exerts anti-inflammatory effects and can protect larvae from death. Collectively, the observation interval in this experiment is reasonably and effectively. 3. After Dr. Malacotrichau2019s reminder, we checked the original data and used GraphPad Prism to plot again. Unfortunately, there were some problems in the preparation of data plot. For example, the survival time of the live larvae should be 72 h instead of 12 h or 20 h in LPS+DEX group. If it is recorded as 12 or 20, it means that the experiment stopped at 12 or 20 h. Therefore, we corrected the figure. The original (A B) and corrected (C D) figures were showed as below. This mistake would not affect the scientific conclusions of this study. We sincerely apologize for this unintentional error and appreciate you for pointing it out!  , 
 #8 1. That certainly explains the irregular time intervals, but raises a brand new question: if you could subject the organisms to  72 h continuous observation , why not do that for all organisms? 2. Furthermore, that doesn t fully explain why exactly one individual at a time would die in all of the other tests. LPS+LGS-L, for example has half of all individuals die, but again never more than 1 per interval; if we assume death rate is Poisson distributed, that seems really unlikely to occur. Admittedly, I can t comment on the aspects you describe in point 2; but from a statistics point of view that does seem unlikely. 3. Your data has again changed. A does not produce B. And how did you delete so much data from C to produce A? And you still haven t discussed how you went from n=50 to n=10. Every time you  check your data your data changes. , 
 #9 Dear Dr. Decaspila, thank you for your comments. 1. The observing and recording time points in all groups are the same as LPS group which is determined by pre-experiments. Similar references are shown as follows: [1] Wynd B M, Watson C J, Patil K, et al. A Dynamic Anesthesia System for Long-Term Imaging in Adult Zebrafish[J]. Zebrafish. 2017, 14(1): 1-7. [2] Zaccaria E, Cao R, Wells J M, et al. A Zebrafish Larval Model to Assess Virulence of Porcine Streptococcus suis Strains[J]. PLOS ONE. 2016, 11(3): e151623. [3] Mordes D A, Morrison B M, Ament X H, et al. Absence of Survival and Motor Deficits in 500 Repeat C9ORF72 BAC Mice[J]. Neuron. 2020, 108(4): 775-783. [4] Ojanen M J T, Uusi-Mu00e4kelu00e4 M I E, Harjula S E, et al. Intelectin 3 is dispensable for resistance against a mycobacterial infection in zebrafish (Danio rerio)[J]. Scientific Reports. 2019, 9(1). 2. In experimental process, we also notice that only one larva or none was dead in drug-treated groups. We speculated that this phenomenon may be cause by the anti-inflammatory effects of dexamethasone and LGS. 3. A can produce B by using Prism software. We didnu2019t delete any data from C to produce A. The detailed reason we have explained in the last reply. We also explained that the group is listed at n=10, but not n=50, previously. ,