Competition and Homeostasis of Excitatory and Inhibitory Connectivity in the Adult Mouse Visual Cortex

Cerebral Cortex, October 2015;25: 3713–3722 doi: 10.1093/cercor/bhu245 Advance Access Publication Date: 14 October 2014 ORIGINAL ARTICLE ORIGINAL ARTICLE Competition and Homeostasis of Excitatory and Inhibitory Connectivity in the Adult Mouse Visual Cortex Department of Molecular Visual Plasticity, The Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Sciences (KNAW), Amsterdam 1105, The Netherlands Address correspondence to Dr Christiaan N. Levelt, Netherlands Institute for Neuroscience, Meibergdreef 47, 1105 BA Amsterdam, The Netherlands. Email: Abstract During cortical development, synaptic competition regulates the formation and adjustment of neuronal connectivity. It is unknown whether synaptic competition remains active in the adult brain and how inhibitory neurons participate in this process. Using morphological and electrophysiological measurements, we show that expressing a dominant-negative form of the TrkB receptor (TrkB.T1) in the majority of pyramidal neurons in the adult visual cortex does not affect excitatory synapse densities. This is in stark contrast to the previously reported loss of excitatory input which occurs if the exact same transgene is expressed in sparse neurons at the same age. This indicates that synaptic competition remains active in adulthood. Additionally, we show that interneurons not expressing the TrkB.T1 transgene may have a competitive advantage and obtain more excitatory synapses when most neighboring pyramidal neurons do express the transgene. Finally, we demonstrate that inhibitory synapses onto pyramidal neurons are reduced when TrkB signaling is interfered with in most pyramidal neurons but not when few pyramidal neurons have this deficit. This adjustment of inhibitory innervation is therefore not a cell-autonomous consequence of decreased TrkB signaling but more likely a homeostatic mechanism compensating for activity changes at the population level. Key words: BDNF, cell autonomous, inhibition, parvalbumin, TrkB Introduction The brain has a strong capacity to learn and adapt to the environment. This adaptation is mainly mediated by the continuous gain, loss, and functional adjustment of synapses based on neuronal activity patterns. Synaptic competition, whereby inputs that are more capable of activating the postsynaptic neuron are strengthened while those that are less capable are lost, is crucial for such adaptation in the developing nervous system. A classic example is ocular dominance plasticity in the primary visual cortex (V1) where monocular deprivation during development causes retraction of thalamic projections serving the closed eye and expansion of those serving the open eye (Hubel et al. 1977). This competitive process operates through 2 separate mechanisms. Correlation-based plasticity weakens the deprived eye responses, while homeostatic synaptic scaling normalizes total postsynaptic drive by increasing the responses to both eyes (Frenkel and Bear 2004; Mrsic-Flogel et al. 2007; Kaneko, Stellwagen et al. 2008; Kaneko, Hanover et al. 2008; Kaneko et al. 2012). Besides presynaptic competition for postsynaptic targets, postsynaptic competition for presynaptic inputs also occurs in the cortex. When expression of neuroligin-1 (Kwon et al. 2012) © The Author 2014. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact M. Hadi Saiepour, Sridhara Chakravarthy, Rogier Min, and Christiaan N. Levelt 3714 | Cerebral Cortex, 2015, Vol. 25, No. 10 Materials and Methods Transgenic Mice We made use of mice carrying a cre-dependent transgene driving expression of either the TrkB.T1 receptor fused to the enhanced green fluorescent protein (EGFP) (mouse line TLT-817) or a membrane associated EGFP-F-transgene (mouse line TLG-498) under the control of the Thy-1 promoter. For sparse expression, these lines were crossed to the CaMKIIα-cre-transgenic mouse line Cre 3487. All the above-mentioned lines were described previously (Chakravarthy et al. 2006, 2008). Broad expression of the TrkB. T1-EGFP transgene in most cortical pyramidal neurons was achieved by crossing the TLT-817 line to G35-3 cre mice which express cre under the control of the KA1 promoter (Sawtell et al. 2003; Heimel et al. 2010). Mice were crossed to C57BL/6JOlaHsd background for at least 6 generations. We will refer to the broadly expressing TLT-817+G35-3-cre+ mice as “TrkB.T1-broad,” and to the sparsely expressing TLT-817+Cre-3487+ mice as “TrkB.T1sparse.” TLG-498+Cre-3487+ double-positive mice are referred to as “EGFP-sparse.” All experiments were approved by the institutional animal care and use committee of the Royal Netherlands Academy of Arts and Sciences. in phosphate buffered saline (PBS). Brains were dissected, postfixed for 2 h, and subsequently stored at 4°C in PBS. Coronal sections of 50 µm were made using a vibratome (Leica VT1000S, Leica Microsystems, Wetzlar, Germany). Free-floating sections were double-stained using mouse anti-NeuN or anti-GFP antibodies (1:500, Chemicon) and rabbit anti-parvalbumin (PV) (1:1000, Swant) followed by Alexa 488-conjugated goat antimouse antibodies (1:500, Molecular Probes) and Alexa 568 conjugated goat anti-rabbit antibodies (1:500, Molecular Probes). Diolistics DiI-coated Tefzel tubing was prepared as described earlier (Chakravarthy et al. 2006). Briefly, a mixture of 0.15 mg DiI (Molecular Probes), 50 μL methylene chloride, and 12 mg of 1.1 μm tungsten particles (Bio-Rad, Veenendaal, the Netherlands) was spread on a glass slide and air-dried. Subsequent to resuspension in 1 mL distilled water and sonication, the mixture was sucked into Tefzel tubing (Bio-Rad). The suspension was withdrawn after 2 min and nitrogen gas was passed to dry the tube. The tube was cut into 13-mm pieces and was used in a Helios Gene Gun (BioRad) to shoot 50-μm coronal sections ( prepared as described for immunohistochemistry) at 100 psi through a membrane filter of 3-μm pore size and 8 × 105 pores/cm2 (Corning, Acton, MA, USA). Sections were stored in PBS at room temperature for least 12 h to ensure good filling of the labeled neurons. Confocal Microscopy DiI-labeled neurons from layer II/III of V1 were imaged using a Carl Zeiss CLSM 510 Meta confocal microscope (Zeiss, Göttingen, Germany) with HeNe (543 nm) laser. Only those DiI-labeled neurons that were also positive for TrkB.T1 were imaged in TrkB.T1broad mice. In the littermate controls, all DiI-labeled pyramidal neurons from layer II/III were imaged. The dendritic segments after the first branch point of basal dendrites were imaged with a ×63 oil-immersion objective and at an optical zoom of ×2.5. Each image was a stack of 50 planes with (...truncated)