Patch clamp analysis of synaptic circuitry in the superior colliculus

Abstract

Background: Patch-clamp recording in brain slices is a powerful method for investigating the neural circuitry of the central nervous system. Applying this approach to a visuomotor center, the superior colliculus, has allowed us to address the fundamental question of how the visual system generates the motor command signals that initiate orienting movements of the head and eyes. Methods: We used the whole-cell patch-clamp method to examine intrinsic synaptic circuitry within the superior colliculus of tree shrews and rats. We first analyzed the synaptic transmission between the layers of the superior colliculus by electrically stimulating the superficial, or visual, layer, which encodes the location of objects in the visual field; and simultaneously recording postsynaptic responses of the intermediate, or motor, layer cells that command eye-orienting movements. We then used UV "photostimulation" to uncage the neurotransmitter glutamate for the analysis of local synaptic interactions within the intermediate layer that may contribute to the generation of the command signals. Results: Stimulating the visual layer evoked inward postsynaptic currents in intermediate layer neurons. Increasing stimulus intensity increased the magnitude of these synaptic responses, indicating that a population of presynaptic inputs converge upon individual presaccadic neurons. The postsynaptic currents evoked by visual layer stimulation were excitatory because they reversed at a postsynaptic membrane potential of approximately 0 mV and were capable of evoking action potentials. An array of stimulating electrodes allowed us to investigate how the responses of the intermediate layer neurons vary according to the position of the stimulus within the visual layer. The largest responses were evoked when the stimulating electrode was aligned directly above a given intermediate layer neuron, indicating a columnar organization of synaptic projections from the visual to the motor layer. One remarkable feature of these postsynaptic responses is that they could last more than 100 times longer than the stimulus applied to the visual layer. These postsynaptic responses are sufficient in duration to generated the prolonged bursts of action potentials that command saccades. This result suggested that one function of superior colliculus circuitry is to enhance and prolong signals that are relayed from the visual to the motor layer. Recording in the superficial layer showed that responses of most of the neurons were transient and can not be the sole presynaptic source of input to the command cells in the motor layer. Photostimulation of individual neurons within the intermediate layer produced substantial excitatory synaptic responses in neighboring command cells. This indicates strong local excitatory connections are present within the motor layers that could be responsible for prolonging the postsynaptic responses of the command cells. Conclusions: Our results demonstrate the presence of a powerful excitatory projection from the visual to the motor layer of the superior colliculus. The columnar organization of the projection suggests that the connections between the layers link corresponding regions of a spatially organized visual map in the superficial layer with a map of the vectors of orienting movements in the intermediate layer. The presence of this projection within the collicular slices suggests that circuitry within the superior colliculus plays a significant role in the generation of visually guided saccades. The long duration of the intermediate layer responses to superficial layer stimulation suggests that this intracollicular circuitry may also strengthen and prolong transient sensory signals into the prolonged bursts of action potentials that have been shown in the awake primate to command saccades. Recording from cells in the superficial layer suggests that the responses of most of the superficial cells themselves are transient and thus cannot be solely responsible for the prolonged bursts in the intermediate layer. Instead, circuits intervening between the visual and motor cells must prolong the transient responses of the visual cells. In support of this idea, photostimulation demonstrates that powerful excitatory circuits exist locally within the intermediate layer that may help transform the transient visual signals arriving from the superficial layer into the prolonged bursts of action potentials that command saccades. Further exploitation of the advantages of high-resolution patch-clamp recordings promises to uncover more details about the functional circuitry in the superior colliculus.