Defining cortical frequency tuning with recurrent excitatory circuitry

Although many aspects of representation/processing function of neurons in the recipient layers of cortex appear to reflect converging thalamocortical inputs1–5, the functional role and the underlying pattern of thalamocortical and, in particular, intracortical excitatory inputs remain unsolved6–9. Extensive efforts have been made to understand the thalamocortical contribution to cortical responses. These previous studies can be mostly categorized into two types: 1) directly compare the response properties between simultaneously recorded neurons in the thalamus and cortex 1,10–12; and 2) isolate thalamocortical input by preventing spiking of cortical neurons2,13–17. The first type of studies mostly used extracellular recordings, and identified putatively connected thalamic and cortical units based on temporal correlation between their spikes. This approach provides information on the tuning properties of individual thalamic and cortical neurons as well as the nature of connection between them. A recent study in the somatosensory cortex5, by pairing extracellular recording of thalamic neurons with intracellular recording of cortical cells, suggests that cortical neurons receive a number of weak but synchronously activated thalamic inputs, which exhibit similar tuning properties as the recorded cortical neuron. However, since the pattern underlying divergent output connections made by a single thalamic neuron, or convergent thalamic inputs made on a single cortical neuron, remains largely unknown, it is difficult to determine the respective functional roles of thalamocortical and intracortical inputs. The second approach depends on an effective silencing of the cortex without affecting thalamocortical transmission. Three methods have been previously used to silence the cortex: 1) cortical application of muscimol, an agonist of GABAA receptors, to prevent neuronal spiking13–15; 2) cooling the cortex (4–14 °C) to block spike generation in neurons2,17,18; 3) electrical stimulation of the cortex to produce a long inhibition widow (>100 ms) following excitation, during which spikes cannot be generated16. However, all these methods are expected to have impacts on thalamocortical presynaptic transmission. Electrical stimulation can result in complex presynaptic effects such as short-term depression or facilitation. Although the mechanism underlying cooling-induced action potential block is not yet clear, it is likely that both the action potential spread in axons and presynaptic vesicle release will also be affected by a drastic temperature decrease. Microinjection, iontophoresis or perfusion of muscimol have more often been applied to silence intracortical connections 13–15. It was assumed that muscimol was a highly specific agonist to GABAA receptors. However this view has been challenged by recent findings that muscimol at a relatively low concentration can already activate GABAB receptors, and reduce synaptic transmission through presynaptic GABAB receptors 19. In this work, we developed a new pharmacological method to silence the cortex. By simultaneously blocking GABAB receptors with a specific antagonist, we could largely prevent the non-specific effect of muscimol on presynaptic transmission. By applying in vivo whole-cell voltage-clamp recording in the rat primary auditory cortex (A1), we examined tone-evoked synaptic responses in layer 4 neurons before and after local cortical silencing. We found that thalamocortical inputs determine the area of synaptic frequency-intensity tonal receptive field (TRF), while intracortical excitatory inputs largely define the frequency tuning by selectively amplifying responses at preferred frequencies of the cortical cell.