Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

PKCα integrates spatiotemporally distinct Ca2+ and autocrine BDNF signaling to facilitate synaptic plasticity

Abstract

The protein kinase C (PKC) enzymes have long been established as critical for synaptic plasticity. However, it is unknown whether Ca2+-dependent PKC isozymes are activated in dendritic spines during plasticity and, if so, how this synaptic activity is encoded by PKC. Here, using newly developed, isozyme-specific sensors, we demonstrate that classical isozymes are activated to varying degrees and with distinct kinetics. PKCα is activated robustly and rapidly in stimulated spines and is the only isozyme required for structural plasticity. This specificity depends on a PDZ-binding motif present only in PKCα. The activation of PKCα during plasticity requires both NMDA receptor Ca2+ flux and autocrine brain-derived neurotrophic factor (BDNF)–TrkB signaling, two pathways that differ vastly in their spatiotemporal scales of signaling. Our results suggest that, by integrating these signals, PKCα combines a measure of recent, nearby synaptic plasticity with local synaptic input, enabling complex cellular computations such as heterosynaptic facilitation of plasticity necessary for efficient hippocampus-dependent learning.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Development of FLIM-FRET sensors for isozyme-specific PKC activity.
Fig. 2: Structural plasticity induces robust, compartmentalized and rapid PKCα activity.
Fig. 3: Classical isozyme PKCα, but not PKCβ or PKCγ, regulates structural plasticity.
Fig. 4: Isozyme-specific PKCα activity and function is defined by a C-terminal PDZ-binding motif.
Fig. 5: Functional and behavioral characterization of plasticity in PKCα KO animals.
Fig. 6: NMDAR and BDNF–TrkB activation converge to activate PKCα.
Fig. 7: Integration of multiple upstream signals by PKCα induces heterosynaptically facilitated plasticity.

Similar content being viewed by others

References

  1. Nicoll, R. A. & Roche, K. W. Long-term potentiation: peeling the onion. Neuropharmacology 74, 18–22 (2013).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  2. Nishiyama, J. & Yasuda, R. Biochemical computation for spine structural plasticity. Neuron 87, 63–75 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  3. Hulme, S. R., Jones, O. D. & Abraham, W. C. Emerging roles of metaplasticity in behaviour and disease. Trends Neurosci. 36, 353–362 (2013).

    Article  PubMed  CAS  Google Scholar 

  4. Harvey, C. D. & Svoboda, K. Locally dynamic synaptic learning rules in pyramidal neuron dendrites. Nature 450, 1195–1200 (2007).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  5. Govindarajan, A., Israely, I., Huang, S. Y. & Tonegawa, S. The dendritic branch is the preferred integrative unit for protein synthesis-dependent LTP. Neuron 69, 132–146 (2011).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  6. Hedrick, N. G. et al. Rho GTPase complementation underlies BDNF-dependent homo- and heterosynaptic plasticity. Nature 538, 104–108 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  7. Makino, H. & Malinow, R. Compartmentalized versus global synaptic plasticity on dendrites controlled by experience. Neuron 72, 1001–1011 (2011).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  8. Fu, M., Yu, X., Lu, J. & Zuo, Y. Repetitive motor learning induces coordinated formation of clustered dendritic spines in vivo. Nature 483, 92–95 (2012).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  9. Takahashi, N. et al. Locally synchronized synaptic inputs. Science 335, 353–356 (2012).

    Article  PubMed  CAS  Google Scholar 

  10. Olds, J. L., Anderson, M. L., McPhie, D. L., Staten, L. D. & Alkon, D. L. Imaging of memory-specific changes in the distribution of protein kinase C in the hippocampus. Science 245, 866–869 (1989).

    Article  PubMed  CAS  Google Scholar 

  11. Pastalkova, E. et al. Storage of spatial information by the maintenance mechanism of LTP. Science 313, 1141–1144 (2006).

    Article  PubMed  CAS  Google Scholar 

  12. Hongpaisan, J. & Alkon, D. L. A structural basis for enhancement of long-term associative memory in single dendritic spines regulated by PKC. Proc. Natl Acad. Sci. USA 104, 19571–19576 (2007).

    Article  PubMed  Google Scholar 

  13. Malinow, R., Schulman, H. & Tsien, R. W. Inhibition of postsynaptic PKC or CaMKII blocks induction but not expression of LTP. Science 245, 862–866 (1989).

    Article  PubMed  CAS  Google Scholar 

  14. Hu, G. Y. et al. Protein kinase C injection into hippocampal pyramidal cells elicits features of long term potentiation. Nature 328, 426–429 (1987).

    Article  PubMed  CAS  Google Scholar 

  15. Malinow, R., Madison, D. V. & Tsien, R. W. Persistent protein kinase activity underlying long-term potentiation. Nature 335, 820–824 (1988).

    Article  PubMed  CAS  Google Scholar 

  16. Routtenberg, A. et al. Phorbol ester promotes growth of synaptic plasticity. Brain Res. 378, 374–378 (1986).

    Article  PubMed  CAS  Google Scholar 

  17. Huang, F. L., Yoshida, Y., Nakabayashi, H., Young, W. S. III & Huang, K. P. Immunocytochemical localization of protein kinase C isozymes in rat brain. J. Neurosci. 8, 4734–4744 (1988).

    Article  PubMed  CAS  Google Scholar 

  18. Clark, E. A., Leach, K. L., Trojanowski, J. Q. & Lee, V. M. Characterization and differential distribution of the three major human protein kinase C isozymes (PKC alpha, PKC beta, and PKC gamma) of the central nervous system in normal and Alzheimer’s disease brains. Lab. Invest. 64, 35–44 (1991).

    PubMed  CAS  Google Scholar 

  19. Kose, A., Ito, A., Saito, N. & Tanaka, C. Electron microscopic localization of gamma- and beta II-subspecies of protein kinase C in rat hippocampus. Brain Res. 518, 209–217 (1990).

    Article  PubMed  CAS  Google Scholar 

  20. Ito, A. et al. Immunocytochemical localization of the alpha subspecies of protein kinase C in rat brain. Proc. Natl Acad. Sci. USA 87, 3195–3199 (1990).

    Article  PubMed  CAS  Google Scholar 

  21. Schleifenbaum, A., Stier, G., Gasch, A., Sattler, M. & Schultz, C. Genetically encoded FRET probe for PKC activity based on pleckstrin. J. Am. Chem. Soc. 126, 11786–11787 (2004).

    Article  PubMed  CAS  Google Scholar 

  22. Violin, J. D., Zhang, J., Tsien, R. Y. & Newton, A. C. A genetically encoded fluorescent reporter reveals oscillatory phosphorylation by protein kinase C. J. Cell Biol. 161, 899–909 (2003).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  23. Braun, D. C., Garfield, S. H. & Blumberg, P. M. Analysis by fluorescence resonance energy transfer of the interaction between ligands and protein kinase Cδ in the intact cell. J. Biol. Chem. 280, 8164–8171 (2005).

    Article  PubMed  CAS  Google Scholar 

  24. Yasuda, R. Imaging spatiotemporal dynamics of neuronal signaling using fluorescence resonance energy transfer and fluorescence lifetime imaging microscopy. Curr. Opin. Neurobiol. 16, 551–561 (2006).

    Article  PubMed  CAS  Google Scholar 

  25. Steinberg, S. F. Structural basis of protein kinase C isoform function. Physiol. Rev. 88, 1341–1378 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  26. Apolloni, A., Prior, I. A., Lindsay, M., Parton, R. G. & Hancock, J. F. H-ras but not K-ras traffics to the plasma membrane through the exocytic pathway. Mol. Cell. Biol. 20, 2475–2487 (2000).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  27. Hayashi-Takagi, A. et al. Labelling and optical erasure of synaptic memory traces in the motor cortex. Nature 525, 333–338 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  28. Matsuzaki, M., Honkura, N., Ellis-Davies, G. C. & Kasai, H. Structural basis of long-term potentiation in single dendritic spines. Nature 429, 761–766 (2004).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  29. Gschwendt, M. et al. Inhibition of protein kinase C mu by various inhibitors. Differentiation from protein kinase c isoenzymes. FEBS Lett. 392, 77–80 (1996).

    Article  PubMed  CAS  Google Scholar 

  30. Sakai, N. et al. Direct visualization of the translocation of the gamma-subspecies of protein kinase C in living cells using fusion proteins with green fluorescent protein. J. Cell Biol. 139, 1465–1476 (1997).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  31. Wagner, S., Harteneck, C., Hucho, F. & Buchner, K. Analysis of the subcellular distribution of protein kinase Cα using PKC-GFP fusion proteins. Exp. Cell Res. 258, 204–214 (2000).

    Article  PubMed  CAS  Google Scholar 

  32. Feng, X. et al. Visualization of dynamic trafficking of a protein kinase C βII/green fluorescent protein conjugate reveals differences in G protein-coupled receptor activation and desensitization. J. Biol. Chem. 273, 10755–10762 (1998).

    Article  PubMed  CAS  Google Scholar 

  33. Staudinger, J., Lu, J. & Olson, E. N. Specific interaction of the PDZ domain protein PICK1 with the COOH terminus of protein kinase C-α. J. Biol. Chem. 272, 32019–32024 (1997).

    Article  PubMed  CAS  Google Scholar 

  34. Minichiello, L. et al. Mechanism of TrkB-mediated hippocampal long-term potentiation. Neuron 36, 121–137 (2002).

    Article  PubMed  CAS  Google Scholar 

  35. Korte, M. et al. Hippocampal long-term potentiation is impaired in mice lacking brain-derived neurotrophic factor. Proc. Natl Acad. Sci. USA 92, 8856–8860 (1995).

    Article  PubMed  CAS  Google Scholar 

  36. Chen, X. et al. A chemical-genetic approach to studying neurotrophin signaling. Neuron 46, 13–21 (2005).

    Article  PubMed  CAS  Google Scholar 

  37. Harward, S. C. et al. Autocrine BDNF–TrkB signalling within a single dendritic spine. Nature 538, 99–103 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  38. Zhai, S., Ark, E. D., Parra-Bueno, P. & Yasuda, R. Long-distance integration of nuclear ERK signaling triggered by activation of a few dendritic spines. Science 342, 1107–1111 (2013).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  39. Chen, T.-W. et al. Ultrasensitive fluorescent proteins for imaging neuronal activity. Nature 499, 295–300 (2013).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  40. Dana, H. et al. Sensitive red protein calcium indicators for imaging neural activity. Elife 5, e12727 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  41. Murakoshi, H. & Shibata, A. C. E. ShadowY: a dark yellow fluorescent protein for FLIM-based FRET measurement. Sci. Rep. 7, 6791 (2017).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  42. Lee, S. J., Escobedo-Lozoya, Y., Szatmari, E. M. & Yasuda, R. Activation of CaMKII in single dendritic spines during long-term potentiation. Nature 458, 299–304 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  43. Terashima, A. et al. An essential role for PICK1 in NMDA receptor-dependent bidirectional synaptic plasticity. Neuron 57, 872–882 (2008).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  44. Woolfrey, K. M. & Dell’Acqua, M. L. Coordination of protein phosphorylation and dephosphorylation in synaptic plasticity. J. Biol. Chem. 290, 28604–28612 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  45. Weeber, E. J. et al. A role for the beta isoform of protein kinase C in fear conditioning. J. Neurosci. 20, 5906–5914 (2000).

    Article  PubMed  CAS  Google Scholar 

  46. Abeliovich, A. et al. Modified hippocampal long-term potentiation in PKC gamma-mutant mice. Cell 75, 1253–1262 (1993).

    Article  PubMed  CAS  Google Scholar 

  47. Abeliovich, A. et al. PKCγ mutant mice exhibit mild deficits in spatial and contextual learning. Cell 75, 1263–1271 (1993).

    Article  PubMed  CAS  Google Scholar 

  48. Freeley, M., Kelleher, D. & Long, A. Regulation of protein kinase C function by phosphorylation on conserved and non-conserved sites. Cell. Signal. 23, 753–762 (2011).

    Article  PubMed  CAS  Google Scholar 

  49. Hoeffer, C. A. & Klann, E. mTOR signaling: at the crossroads of plasticity, memory and disease. Trends Neurosci. 33, 67–75 (2010).

    Article  PubMed  CAS  Google Scholar 

  50. Kastellakis, G., Cai, D. J., Mednick, S. C., Silva, A. J. & Poirazi, P. Synaptic clustering within dendrites: an emerging theory of memory formation. Prog. Neurobiol. 126, 19–35 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  51. Sacktor, T. C. et al. Persistent activation of the zeta isoform of protein kinase C in the maintenance of long-term potentiation. Proc. Natl Acad. Sci. USA 90, 8342–8346 (1993).

    Article  PubMed  CAS  Google Scholar 

  52. Leitges, M. et al. Immunodeficiency in protein kinase Cβ-deficient mice. Science 273, 788–791 (1996).

    Article  PubMed  CAS  Google Scholar 

  53. Leitges, M. et al. Knockout of PKC alpha enhances insulin signaling through PI3K. Mol. Endocrinol. 16, 847–858 (2002).

    PubMed  Google Scholar 

  54. He, X. P. et al. Conditional deletion of TrkB but not BDNF prevents epileptogenesis in the kindling model. Neuron 43, 31–42 (2004).

    Article  PubMed  CAS  Google Scholar 

  55. Stoppini, L., Buchs, P. A. & Muller, D. A simple method for organotypic cultures of nervous tissue. J. Neurosci. Methods 37, 173–182 (1991).

    Article  PubMed  CAS  Google Scholar 

  56. O’Brien, J. A. & Lummis, S. C. Biolistic transfection of neuronal cultures using a hand-held gene gun. Nat. Protoc. 1, 977–981 (2006).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  57. Murakoshi, H., Wang, H. & Yasuda, R. Local, persistent activation of Rho GTPases during plasticity of single dendritic spines. Nature 472, 100–104 (2011).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  58. Pologruto, T. A., Sabatini, B. L. & Svoboda, K. ScanImage: flexible software for operating laser scanning microscopes. Biomed. Eng. Online 2, 13 (2003).

    Article  PubMed  PubMed Central  Google Scholar 

  59. Smirnov, M.S., Evans, P.R., Garrett, T.R., Yan, L. & Yasuda, R. Automated remote focusing, drift correction, and photostimulation to evaluate structural plasticity in dendritic spines. Preprint at bioRxiv https://doi.org/10.1101/083006 (2016).

  60. Sugar, J. D., Cummings, A. W., Jacobs, B. W. & David, B. Robinson. A free Matlab script for spatial drift correction. Micros. Today 22, 40–47 (2014).

    Article  Google Scholar 

  61. Geusebroek, J. M., Cornelissen, F., Smeulders, A. W. & Geerts, H. Robust autofocusing in microscopy. Cytometry 39, 1–9 (2000).

    Article  PubMed  CAS  Google Scholar 

  62. Laviv, T. et al. Simultaneous dual-color fluorescence lifetime imaging with novel red-shifted fluorescent proteins. Nat. Methods 13, 989–992 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  63. Deacon, R. M. & Rawlins, J. N. T-maze alternation in the rodent. Nat. Protoc. 1, 7–12 (2006).

    Article  PubMed  Google Scholar 

  64. Vorhees, C. V. & Williams, M. T. Morris water maze: procedures for assessing spatial and related forms of learning and memory. Nat. Protoc. 1, 848–858 (2006).

    Article  PubMed  PubMed Central  Google Scholar 

  65. Wenk, G. L. Assessment of spatial memory using the radial arm maze and Morris water maze. Curr. Protoc. Neurosci. 8, 8.5A, https://doi.org/10.1002/0471142301.ns0805as26 (2004).

    Article  Google Scholar 

  66. Pham, J., Cabrera, S. M., Sanchis-Segura, C. & Wood, M. A. Automated scoring of fear-related behavior using EthoVision software. J. Neurosci. Methods 178, 323–326 (2009).

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

We would like to thank A. F. Brantley and the Scripps Florida behavior core, who performed animal behavioral studies; L. Parada (Memorial Sloan Kettering) for Bdnf fl/fl mice; D. Ginty (Harvard) for TrkBF616A mice; M. Dowdy and the MPFI ARC for animal care; members of the Yasuda laboratory; L. Yan; and D. Kloetzer. This work was funded by F32MH101954 (L.A.C.), R01MH080047 (R.Y.), 1DP1NS096787 (R.Y.) and the Max Planck Florida Institute for Neuroscience.

Author information

Authors and Affiliations

Authors

Contributions

Conceptualization: L.A.C., R.Y. Investigation: L.A.C., M.H., J.A.M., P.P.-B., C.M.M. Resources: M.L. Writing: original draft: L.A.C. Writing: review and editing: L.A.C., R.Y. Funding acquisition: L.A.C., R.Y.

Corresponding authors

Correspondence to Lesley A Colgan or Ryohei Yasuda.

Ethics declarations

Competing interests

The authors declare no competing financial or non-financial interests as defined by Nature Research.

Additional information

Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Integrated supplementary information

Supplementary Figure 1 Classical PKC isozyme activity during sLTP.

A,B) Schematic of ITRACKneg (A) and IDOCKneg (B). C,D) Overlay of mean time courses of PKCα, PKCβ, and PKCγ activity as measured by ITRACK (C; n[neurons/spines]: PKCα=32/103 (same as Figure 2D), PKCβ =13/31, PKCγ = 10/27) or IDOCKS (D; n[neurons/spines]: PKCα = 6/28(same as in Figure 2K), PKCβ = 6/24, PKCγ= 8/29). Insets: Quantification of area under the curve (mean and SEM) of PKC isozymes to each glutamate uncaging pulse (as in Figure 2C,L). One way ANOVA with Sidak’s multiple comparison compared to PKCα. E) Average translocation in response to each glutamate uncaging pulse (uncaging triggered average) of ITRACKα (same data as in C), ITRACKα in the absence of glutamate (n[neurons/spines]= 6/15) or the control sensor ITRACKα neg (n[neurons/spines]= 6/19). Mean and SEM (shaded) are shown. F) Average translocation in response to each glutamate uncaging pulse (uncaging triggered average) of IDOCKSα (same data as in D), IDOCKSα in the absence of glutamate (n[neurons/spines]= 5/12) or the control sensor IDOCKSα neg (n[neurons/spines]= 7/20). Mean and SEM (shaded) are shown.

Supplementary Figure 2 PKCα activity is not modulated by sensor design or expression.

A) Change in ITRACKα activity as a function of the expression level of the ITRACKα donor measured by intensity of eGFP at the primary dendrite (n[neurons/spines]= 19/55) and compared to standards of purified eGFP of known concentration. Pearson’s correlation coefficient (r2) indicated. B) Change in ITRACKα activity as a function of the relative expression level of the donor to the acceptor as measured by intensity of eGFP and mCh excited with 920nm at the primary dendrite (n[neurons/spines] = 19/55). Pearson’s correlation coefficient (r2) indicated. C) Change in basal lifetime of eGFP (n[neurons/spines] 1/63), ITRACKα (n[neurons/spines]=1/60) or IDOCKSα (n[neurons/spines]=1/47) as a function of spine size. Pearson’s correlation coefficient (r2) indicated. D) Mean basal lifetime of eGFP (n: spines=63, dendrites=16), ITRACKα (n: spines=60, dendrites=15) or IDOCKSα (n: spines=47, dendrites=15) measured in spines or dendrites of a single neuron. Two-tailed, unpaired t-test with Welch’s correction not significant. E) Average timecourse of fluorescence decay in spines after photoactivation of paGFP (τ =0.91, n[neurons/spines]=4/78) or paGFP-PKCα (τ =2.19, n[neurons/spines]=3/40). F) Average timecourse of fluorescence decay in spines after photoactivation paGFP-PKCα (same as in E) or paGFP-PKCα and ITRACK acceptor construct mCh-CAAX (n[neurons/spines]=3/48) or paGFP-PKCα and IDOCKS acceptor construct 2mCh-PSα (n[neurons/spines]=2/29). G) Quantification of mean sustained sLTP (25-30 min) in neurons expressing eGFP (n[neurons/spines] = 7/11), ITRACKα (n[neurons/spines] = 5/7), or IDOCKSα (n[neurons/spines] = 5/8). One way ANOVA non-significant. H) Average change in PKCα activity to each uncaging pulse (uncaging triggered average) measured with ITRACKα (same as Figure 3D) or a modified ITRACKα in which the acceptor fluorophore was targeted to the membrane with an H-Ras derived CAAX domain (n[neurons/spines] = 3/12). Two way ANOVA not significant by sensor design (F (1, 1872) = 2.769, p=0.0963). For correlation analysis (panels A-C) lines represent linear regression and 95% confidence intervals (dotted). For averaged data (panels D-H) mean and SEM are shown.

Supplementary Figure 3 Requirement of PKC isozymes for sLTP.

A) Mean + SEM sustained sLTP (25-30 min) in the presence of Gö6983 added 10 minutes (n [neurons/spines] = 5/5) or 45 minutes (n [neurons/spines] = 7/7) before the induction of plasticity. Unpaired two-tailed t- test not significant. B) Expression level of rescue constructs eGFP-tagged PKCα, PKCβ, and PKCγ (for experiments in Figure 3E, F; n[neurons/spines]: KO+PKCα = 9/21, KO+PKCβ = 6/16, KO+PKCγ = 6/16). Line represents median value of expression and error is 95% CI of median. C) Overlay of mean time courses of sLTP and SEM of neurons from WT and PKCα KO littermate mice (shown in Figure 3E; n[neurons/spines]: WT = 10/21, KO = 12/26) and WT and PKC α,β,γ TKO mice (as quantified in Figure 3G; n[neurons/spines]: WT=8/21, TKO=7/18). Two way ANOVA significant by genotype (F (3, 1480) = 44.05, p<0.0001). D) Average time courses of sLTP and SEM in eGFP expressing neurons from WT and TKO mice and neurons from TKO mice overexpressing eGFP-PKCα (as quantified in Figure 3G; n[neurons/spines]: WT=8/21, TKO=7/18, TKO +PKCα=7/17). Two way ANOVA significant by genotype (F (1, 560) = 51.08, p<0.0001).

Supplementary Figure 4 Behaviors that were not affected in PKCα KO animals.

Performance of male PKCα KO (n[animals] = 13) and WT (n[animals] = 15) littermate mice in open field (A, Mean and SEM shown, two-tailed unpaired t test), spontaneous alternation with and without delay (B, Mean shown, two-sided Fisher’s exact test) and hot plate test (C, Mean and SEM shown, two-tailed unpaired t-test).

Supplementary Figure 5 Upstream mechanisms of PKCα activation.

A) Quantification of mean sustained structural plasticity of neurons treated with indicated pharmacologic agents. One way ANOVA with Dunnett’s multiple comparison test against controls (n[neurons/spines]: CTL=11/13, APV (50 µM) = 5/9, Edel (50 µM) = 6/13, CTL= 13/15, MCPG (250 µM) = 5/9, NPS (20 µM) = 8/13, CTL=9/9, Veh= 4/7, 1NMPP1 (1 µM) = 7/12. B) Average change in PKCα activity (measured with IDOCKSα) in response to each glutamate uncaging pulse (uncaging triggered average) before (n[neurons/spines] = 5/14) and after edelfosine application (50 µM, n[neurons/spines] = 5/17). Two way ANOVA significant by drug (F (1, 464) = 30.94, p<0.0001). C) Average change in PKCα activity (measured by ITRACKα) in response to each glutamate uncaging pulse (uncaging triggered average) before (n[neurons/spines] = 7/24) and after U73122 application (10 µM, n[neurons/spines]= 3/9) or application of the inactive analog U73343 (10 µM, n[neurons/spines]= 4/13). Two way ANOVA by drug is significant by drug (F (1, 31) = 10.54, p=0.0028). U73343 is not significant compared to CTL (F (1, 35) = 0.5582, p=0.46). D) Average change in PKCα activity (measured by ITRACKα) in response to each glutamate uncaging pulse (uncaging triggered average) before (n[neurons/spines] = 9/25) and after NPS application (20 µM, n[neurons/spines]= 9/28). Two way ANOVA by drug is non-significant (F (1, 816) = 0.0009298, p=0.9757). E) Average change in PKCα activity (measured with IDOCKSα) in response to each glutamate uncaging pulse (uncaging triggered average) in neurons from TrkBF616A mice before (n[neurons/spines] = 5/17) and after 1NMPP1 application (1 µM, n[neurons/spines]= 5/18). Two way ANOVA significant by drug (F (1, 528) = 37.84, p<0.0001). For all panels data shown is mean and SEM.

Supplementary Figure 6 Comparison of distance between paired spines and PKCα activation.

PKCα activation (measured by ITRACKα) in individual spines receiving subthreshold stimulation after induction of sLTP in a nearby spine plotted as a function of distance to the nearby spine (same data as in Figure 7 F, G (paired), n [neurons/spines= 8/15). Pearson’s correlation is non-significant (p=0.956, r2=0.0026).

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–6 and Supplementary Note

Reporting Summary

Supplementary Video 1 - PKCα activity in a single spine undergoing structural plasticity.

Video of PKCα activity (Fig. 1b) during the induction of sLTP by two-photon glutamate uncaging. Fluorescence lifetime images of the FRET sensor ITRACKα were acquired at 8 Hz and uncaging pulses were delivered at 0.5 Hz (indicated by arrowheads). Only the first eight uncaging pulses are shown. Fluorescence lifetime lookup table spans 2.65 ns (blue) to 2.3 ns (red), with PKCα activation corresponding to warm (red) colors

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Colgan, L.A., Hu, M., Misler, J.A. et al. PKCα integrates spatiotemporally distinct Ca2+ and autocrine BDNF signaling to facilitate synaptic plasticity. Nat Neurosci 21, 1027–1037 (2018). https://doi.org/10.1038/s41593-018-0184-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41593-018-0184-3

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing