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Multiphoton imaging of genetically encoded calcium indicators is routinely used to report activity from populations of spatially resolved neurons in vivo. However, since the relationship between fluorescence and action potentials (APs) is nonlinear and varies over neurons, quantitatively inferring AP discharge is problematic. To address this we dev...
Multiphoton imaging (MPI) is widely used for recording activity simultaneously from many neurons in superficial cortical layers in vivo. We combined regenerative amplification multiphoton microscopy (RAMM) with genetically encoded calcium indicators to extend MPI of neuronal population activity into layer 5 (L5) of adult mouse somatosensory cortex....
To facilitate automated patch clamp measurements of ion channels in cells, the development of an all-glass Chiptip pipette is reported that may be combined with the previously described Flip-the-Tip technology. A single measurement requires less than 50 cells, and the addition of drugs for screening can be limited to very low volumes down to 1 micr...
A conventional borosilicate glass patch pipette is glued into a plastic jacket, forming the entity of a FlipTip. One or two three-channel modules of recording tip sockets are mounted on a liquid handler platform to take up FlipTips. The tip sockets are connected to preamplifiers (HEKA) and to a suction system. The inner chamber of the tip sockets i...
Patch clamping facilitates directly controlling the voltage of a cell and thus has been acknowledged to be the gold standard for assessing ion-channel physiology in academic and industrial laboratories. The method was developed 30 years ago by (1976), and its various different configurations with glass electrodes have been successfully applied in m...
Striatal spiny projection (SP) neurons control movement initiation by integrating cortical inputs and inhibiting basal ganglia outputs. Central to this control lies a "microcircuit" that consists of a feedback pathway formed by axon collaterals between GABAergic SP neurons and a feedforward pathway from fast spiking (FS) GABAergic interneurons to S...
Up states are prolonged membrane potential depolarizations critical for synaptic integration and action potential generation in cortical and striatal neurons. They commonly result from numerous concurrent synaptic inputs, whereas neurons reside in a down state when synaptic inputs are few. By quantifying the composition, frequency, and amplitude of...
The deep cerebellar nuclei (DCN) constitute the major structures by which the cerebellum forwards its output to the rest of the brain. Although the connectivity of the DCN has been well studied, little is known about the interface-the neurons' soma and dendrites-between the DCN's inputs and outputs. We therefore decided to analyze the neurons' soma...
Striatal inhibition plays an important role in models of cortex-basal ganglia function and is altered in many basal ganglia diseases. The gamma-aminobutyric acid ergic spiny projection neuron comprises >95% of striatal neurons, but despite strong anatomical evidence, the electrophysiological properties and functions of their local axon collaterals...
Classically, three classes of neurons in the cerebellar nuclei (CN), defined by different projection targets and content of transmitters, have been distinguished. However, evidence for different types of neurons based on different intrinsic properties is lacking. The present study reports two types of neurons defined mainly by their intrinsic prope...
The neuronal cell line NG108-15 (180CC15) responds to extracellular stimuli of ATP or UTP with a transient increase in the level of cytosolic Ca2+. Desensitization was investigated by recording single-cell Ca2+ responses induced by consecutive, regularly spaced (100 s intervals), brief pulses of the nucleotides. The two natural ligands of the P2U r...
In single rat glioma cells, the signal transduction process activated by the UTP sensitive purinergic nucleotide receptor was studied by determining [Ca2+]i by Fura-2 fluorescence and measuring pH by BCECF fluorescence to elucidate the control of [Ca2+]i oscillations by intracellular pH. Addition of UTP for long time periods (some min) causes a [Ca...
Thrombin at nanomolar concentrations induces rapid changes in the second messenger Ca2+ in a glial astrocyte-type cell line. Continuous application of the protease thrombin causes regular [Ca2+]i oscillations (amplitude 109 nM, spike length 48 s) which are suppressed by hirudin. Reduction of [Ca2+]ex (from 1.8 mM to 50 microM) reversibly abolishes...
