PIP3 controls synaptic function by maintaining AMPA receptor clustering at the postsynaptic membrane:

Welcome to Mike and Rohit's Blog O' Science!  We are grad students at Duke and noticed a dearth of good neuroscience blogs, and people kept telling Rohit he should start a blog, so here we are.  I'll be posting my thoughts on papers and current research in the synaptic plasticity field, while Rohit will keep us up to date on in vivo imaging.

The first paper I'd like to cover is a recent paper from the Esteban lab about PI3K signaling during LTP.  Of all the multitudinous signaling pathways involved in LTP, the PI3K pathway was only recently discovered (Sanna et al 2002; Man, et al 2003).  Those papers disagreed slightly on the role of PI3K - Sanna believed it was only necessary for maintenance, while Man thought it was necessary for induction - but both showed that inhibition of PI3K signaling impaired LTP.

This paper started by showing that inhibition of PI3K signaling via overexpressiong PH-Grp1 decreased the basal AMPAR EPSCs in CA1 neurons. PH domains bind to phosphoinositides, and PH-Grp1 specifically binds to PIP3.  Overexpression of PH-Grp1 would thus sequester PIP3, and stop it from functioning in the cell.  They confirmed their genetic result pharmacologically using LY294002, a PI3K inhibitor.  To show that the current decrease was synapse specific, they bath applied AMPA, and showed that the current was the same, and thus number of surface AMPAR was the same.  So far so good.

Next they tried to induce LTP in neurons overexpressing PH-Grp1, and were unable to do so.  Still cool.

To confirm that the decrease in AMPAR currents was due to a decrease in synaptic but not extrasynaptic AMPAR, they transfected neurons with GFP-GluR2 and PH-Grp1.  Here they found that without Grp1, GFP-GluR2 was expressed in both the spine and dendrite; with Grp1, however, GFP-GluR2 shifted more towards the spine.  I originally thought this contradicted other lab's findings, but in verifying this I found that GFP-GluR2 does not have a strong spine bias, while SEP-GluR2 (surface) is punctate in the spine, presumably due to GluR2-containing endosomes in the dendrite (Kopec et. al. 2006).  Still, this result contradicts their earlier finding that Grp1 decreased synaptic but not extrasynaptic AMPAR number.

Since they saw changes in the subcellular distribution of AMPAR when Grp was expressed, they looked at the expression pattern of PSD-95, an integral anchoring protein at the synapse.  They found that in contrast to GluR2, PSD-95 is punctate in control conditions, and loses its punctility after Grp expression.  This contradicts the previous figure, but is in line with the decrease of synaptic AMPAR currents.  To see if this PSD-95 expression pattern changed AMPAR mobility, they performed FRAP on SEP-GluR2, and found that the mobile fraction increased following Grp expression, presumably due to lack of anchoring.  I am usually quite skeptical of FRAP experiments due to the high variability I have encountered doing them myself, and the large disagreement between labs on time constants and mobile fractions.  The FRAP performed here particularly stands out for its bizarrely low mobile fraction and tau recovery (tau of 5 minutes compared to 1-2 minutes in Ashby et. al., 2006 and Makino et. al., 2009).

To try to reconcile the previous two figures, they performed immuno EM to see where in the PSD the AMPAR were located.  Here they found that the AMPAR redistributed from the PSD proper to the perisynaptic region.  While EM has amazing resolution, I am skeptical that a shift of 80 nm in a subset of proteins can be precisely observed.  This finding further does not really explain how the AMPAR are being anchored near the synapse, since PSD-95 binding to TARPs (and thereby, presumably to AMPAR) is intact .

In the discussion the authors mentioned that PIP3, the PI3K end product, is involved in cell polarity, and is present at the tips of branching neurites. They hypothesized that it was playing a similar role in the synapse, which is also a highly polarized compartment.

In the end I found this a frustrating, if interesting paper.  PI3K signaling is an important, if ignored, player in LTP.  However, the contradictory nature of their results is dissatisfying.  I have a hard time understanding how PIP3 simultaneously bring AMPAR into the spine, while reducing AMPAR currents and reducing PSD-95 puncta.  The most intriguing part of this paper, in fact, is the redistribution of PSD-95, which makes me believe future research in PIP3 signaling should focus there, instead of on AMPAR trafficking.

Ashby, M. C., Maier, S. R., Nishimune, A., & Henley, J. M. (2006) Lateral Diffusion Drives Constitutive Exchange of AMPA Receptors at Dendritic Spines and Is Regulated by Spine Morphology. J Neuroscience 26, 7046-7055.

Kopec, C. D., Li, B., Wei, W., Boehm, J., & Malinow, R. (2006) Glutamate Receptor Exocytosis and Spine Enlargement during Chemically Induced Long-Term Potentiation. J Neuroscience 26, 2000-2009.

Makino, H. & Malinow, R. (2009) AMPA Receptor Incorporation into Synapses during LTP: The Role of Lateral Movement and Exocytosis. Neuron 64, 381-390.

Man, H.-Y., Wang, Q., Lu, W.-Y., Ju, W., Ahmadian, G., Liu, L., D'Souza, S., Wong, T. P., Taghibiglou, C., Lu, J., et al. (2003) Activation of PI3-Kinase Is Required for AMPA Receptor Insertion during LTP of mEPSCs in Cultured Hippocampal Neurons. Neuron 38, 611-624.

Sanna, P. P., Cammalleri, M., Berton, F., Simpson, C., Lutjens, R., Bloom, F. E., & Francesconi, W. (2002) Phosphatidylinositol 3-Kinase Is Required for the Expression But Not for the Induction or the Maintenance of Long-Term Potentiation in the Hippocampal CA1 Region. J Neurosci 22, 3359-3365.