Cortical maps, consisting of orderly arrangements of functional columns, are a hallmark of the organization of the cerebral cortex. However, the microorganization of cortical maps at the level of single neurons is not known, mainly because of the limitations of available mapping techniques. Here, we used bulk loading of Ca(2+) indicators combined with two-photon microscopy to image the activity of multiple single neurons in layer (L) 2/3 of the mouse barrel cortex in vivo. We developed methods that reliably detect single action potentials in approximately half of the imaged neurons in L2/3. This allowed us to measure the spiking probability following whisker deflection and thus map the whisker selectivity for multiple neurons with known spatial relationships. At the level of neuronal populations, the whisker map varied smoothly across the surface of the cortex, within and between the barrels. However, the whisker selectivity of individual neurons recorded simultaneously differed greatly, even for nearest neighbors. Trial-to-trial correlations between pairs of neurons were high over distances spanning multiple cortical columns. Our data suggest that the response properties of individual neurons are shaped by highly specific subcolumnar circuits and the momentary intrinsic state of the neocortex.

The functions of cortical areas depend on their inputs and outputs, but the detailed circuits made by long-range projections are unknown. We show that the light-gated channel channelrhodopsin-2 (ChR2) is delivered to axons in pyramidal neurons in vivo. In brain slices from ChR2-expressing mice, photostimulation of ChR2-positive axons can be transduced reliably into single action potentials. Combining photostimulation with whole-cell recordings of synaptic currents makes it possible to map circuits between presynaptic neurons, defined by ChR2 expression, and postsynaptic neurons, defined by targeted patching. We applied this technique, ChR2-assisted circuit mapping (CRACM), to map long-range callosal projections from layer (L) 2/3 of the somatosensory cortex. L2/3 axons connect with neurons in L5, L2/3 and L6, but not L4, in both ipsilateral and contralateral cortex. In both hemispheres the L2/3-to-L5 projection is stronger than the L2/3-to-L2/3 projection. Our results suggest that laminar specificity may be identical for local and long-range cortical projections.

Inducible and reversible silencing of selected neurons in vivo is critical to understanding the structure and dynamics of brain circuits. We have developed Molecules for Inactivation of Synaptic Transmission (MISTs) that can be genetically targeted to allow the reversible inactivation of neurotransmitter release. MISTs consist of modified presynaptic proteins that interfere with the synaptic vesicle cycle when crosslinked by small molecule "dimerizers." MISTs based on the vesicle proteins VAMP2/Synaptobrevin and Synaptophysin induced rapid ( approximately 10 min) and reversible block of synaptic transmission in cultured neurons and brain slices. In transgenic mice expressing MISTs selectively in Purkinje neurons, administration of dimerizer reduced learning and performance of the rotarod behavior. MISTs allow for specific, inducible, and reversible lesions in neuronal circuits and may provide treatment of disorders associated with neuronal hyperactivity.

Can neuronal morphology predict functional synaptic circuits? In the rat barrel cortex, ’barrels’ and ’septa’ delineate an orderly matrix of cortical columns. Using quantitative laser scanning photostimulation we measured the strength of excitatory projections from layer 4 (L4) and L5A to L2/3 pyramidal cells in barrel- and septum-related columns. From morphological reconstructions of excitatory neurons we computed the geometric circuit predicted by axodendritic overlap. Within most individual projections, functional inputs were predicted by geometry and a single scale factor, the synaptic strength per potential synapse. This factor, however, varied between projections and, in one case, even within a projection, up to 20-fold. Relationships between geometric overlap and synaptic strength thus depend on the laminar and columnar locations of both the pre- and postsynaptic neurons, even for neurons of the same type. A large plasticity potential appears to be incorporated into these circuits, allowing for functional ’tuning’ with fixed axonal and dendritic arbor geometry.

Current thinking about long-term memory in the cortex is focused on changes in the strengths of connections between neurons. But ongoing structural plasticity in the adult brain, including synapse formation/elimination and remodelling of axons and dendrites, suggests that memory could also depend on learning-induced changes in the cortical 'wiring diagram'. Given that the cortex is sparsely connected, wiring plasticity could provide a substantial boost in storage capacity, although at a cost of more elaborate biological machinery and slower learning.