We made whole-cell recordings from layer 2/3 pyramidal cells in the somatosensory and visual

cortex to monitor somatic voltage changes, and activated an increasing number of synapses with two-photon learn more glutamate uncaging (Gasparini and Magee, 2006, Losonczy and Magee, 2006 and Matsuzaki et al., 2001). We selected seven spines distributed over a region of ∼20–30 μm (Figure 1A), and recorded somatic EPSPs in response to the activation of one to all seven synapses (with a 1 ms interval between stimulation of each synapse). We found that the EPSP peak increased with the number of activated synapses, closely following a sigmoidal function that greatly deviated from the linear summation of each individual synapse (Figure 1B). We then tested different regions between the tip and the branch point of single branches, and analyzed how this function varied with location. Distal synapses had a much steeper function than proximal synapses (fraction of maximum per input

for tip: 0.48 ± 0.09, middle: 0.24 ± 0.06, LGK-974 in vivo base: 0.11 ± 0.01; n = 9, p = 0.004, ANOVA), which was also shifted to the left (number of inputs at half of the maximum for tip: 2.1 ± 0.4; middle: 3.4 ± 0.6; base: 6.1 ± 0.6, p = 0.006, ANOVA; Figures 1C and Figure S1A, available online). The gain of the input-output function increased more than 3-fold from the branch point to the dendritic tip, and was shifted by approximately five inputs (Figure 1D). Furthermore, the EPSP supralinearity increased from 128% ± 6% to 209% ± 16% between the base and the tip of the dendrite (p = 0.015, ANOVA; Figures 1E and S1B). These results were also observed with small unitary gluEPSPs (∼0.25 mV, see Figures S2A–S2D) and show that there is a gradient of nonlinear synaptic integration along individual dendritic branches, in which distal inputs are amplified more Megestrol Acetate strongly than proximal ones. To understand the biophysical

mechanism underlying supralinear integration, we used pharmacology to probe the role of specific dendritic active conductances (Johnston and Narayanan, 2008, Magee, 2000 and Spruston, 2008). Blocking L-type voltage-gated calcium channels (VGCCs) shifted the input-output curve to the right (Figure 2A; number of inputs at half maximum = 215% ± 39% of control; p = 0.032; supralinearity at three inputs = 118% ± 10%; p = 0.0078; n = 6) without significantly affecting the gain (fraction of maximum per input: 115% ± 13% of control; p = 0.33; Figures 2A and 2C). A similar effect was produced by blocking voltage-gated sodium channels (number of inputs at half maximum = 159% ± 15% of control; p = 0.030; supralinearity at three inputs = 127% ± 15%; p = 0.024; n = 4; Figure 2C), as well as by simultaneous VGCC and voltage-gated sodium channel (VGSC) block (Figure S3).