The patch-clamp technique in the study of secretion.

One of the basic cellular functions of virtually every cell type is the exocytotic release of molecules synthesized, stored and packaged into intracellular vesicles or granules. Over decades much effort has been concentrated on elucidating the chain of events leading to exocytosis. Unfortunately, the nature of the process that ultimately induces membrane fusion is not known, nor has it been established definitively whether or not the final steps in the secretory cascade are identical in different cells. Although the fusion between vesicle and plasma membrane has been neatly documented by electron micrographs, it was only recently that the technique of time-resolved membrane capacitance measurement has provided a more detailed insight into mechanistic aspects of exocytosis, both in terms of the fusion event and the steps involved in stimulus-secretion coupling

Cellular mechanisms of auditory processing in the inferior colliculus: An in vivo patch clamp study

In this thesis, different aspects of central auditory processing in the inferior colliculus (IC) of young-adult mice and rats are described. With the “in vivo patch-clamp” technique we investigated the contribution of membrane propertie

Measurement of the cell membrane capacitance and conductance of colonic crypt cells of the rat using the patch clamp technique

Using the patch clamp technique the membrane capacitance and membrane conductance of colonic crypt cells of the rat was measured. The influence of the intracellular agonists Ca++, cAMP and of osmotic changes on the membrane capacitance and conductance was studied.Comment: Diploma thesis, University of Freiburg, Germany (in German

Batch Fabrication of High-Performance Planar Patch-Clamp Devices in Quartz

The success of the patch-clamp technique has driven an effort to create wafer-based patch-clamp platforms. We develop a lithographic/electrochemical processing scheme that generates ultrasmooth, high aspect ratio pores in quartz. These devices achieve gigaohm seals in nearly 80% of trials, with the majority exhibiting seal resistances from 20-80 GΩ, competing with pipette-based patch-clamp measurements

Sodium channel inactivation kinetics of rat sensory and motor nerve fibres and their modulation by glutathione

Na+ channel currents of rat motor and sensory nerve fibres were studied with the patch-clamp technique on enzymatically demyelinated axons. Differences between motor and sensory fibres in multi-channel inactivation kinetics and the gating of late single-channel currents were investigated. In the axon-attached mode, inactivation of multi-channel Na+ currents in sensory axons was best fitted with a single time constant while for motor axons two time constants were needed. Late single-channel currents in sensory axons were characterized by short openings whereas motor axons exhibited additional long single-channel openings. In contrast, in excised, inside-out membrane patches, no differences between motor and sensory fibres were found: in both types of fibre inactivation of multi-channel Na+ currents proceeded with two time constants and late single-channel currents showed short and long openings. After application of the reducing agent glutathione to the cytoplasmic side of excised inside-out patches, inactivation of Na+ currents in both motor and sensory fibres proceeded with a single, fast exponential time constant and late currents appeared with short openings only. These data indicate that the axonal metabolism may contribute to the different inactivation kinetics of Na+ currents in motor and sensory nerve fibres

Measuring activity of the subthalamic nucleus in acute slices using multi electrode arrays

The symptoms of Parkinson’s disease (a.o.: tremor, rigidity) can be suppressed by electrical stimulation of the basal ganglia. The most common target nucleus of this so called Deep Brain Stimulation (DBS) is the subthalamic nucleus (STN). Good clinical results are obtained by the application of pulses of 200 s, 1-3 V amplitude at a constant rate of about 130 Hz. However, the mechanism(s) responsible for the clinical improvements are not yet elucidated.\ud The use of acute brain slices as a model is widely used, despite the inevitable loss of many connections. Accurate (i.e. subthreshold) measurements of single neuron and multiple neuron (up to ~3, for practical reasons) membrane potentials are obtained by patch-clamp technique. We propose to use arrays of microelectrodes in slice recordings of STN. We present here our first results