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Cutting of living hippocampal slices by a highly pressurised water jet (macromingotome)
D Bingmann1, M Wiemann, E J Speckmann
1Institute of Physiology, University-GH Essen, Hufelandstr. 55, 45122 Essen, Germany. dieter.bingmann@uni-essen.de
Abstract:
Living brain slices are usually cut with razor blades, which compress a ca. 50-microm-thick layer of tissue. This results in cell debris and lesioned cells which, e.g. form diffusion barriers between the bath and living neurons underneath, thereby prolonging response times of neurons to drugs in the bath saline and impeding the experimental access to intact neurons. To avoid such drawbacks, a macromingotome was developed which cuts nervous tissue with water jets. Physiological saline under pressures of 100-1800 bar was ejected through nozzles of 35-100 microm to cut 300-500-microm-thick hippocampal slices. Systematic variations of pressure and nozzle diameter revealed best results at 400-600 bar and with nozzle diameters of 60-80 microm. Under these conditions, intact CA1- and CA3-neurons as well as granule cells were detected with infrared microscopy at less than 10 microm underneath the surface of the slice. Superficial neurons with intact fine structures were also seen when the slices were studied by light-microscopy. Intra- and extracellular recordings from superficial neurons showed normal membrane- and full action potentials and the development of stable epileptiform discharges in 0 Mg(2+)-saline. These results indicate that the macromingotome offers an alternative way of cutting slices which may facilitate electrophysiological/neuropharmacological or fluorometric studies on superficial neurons.
Insights
A novel macromingotome uses water jets to cut brain slices, preserving neuron integrity. This method overcomes limitations of traditional cutting, enabling better studies of living neurons.
Area of Science:
- Neuroscience
- Biotechnology
- Surgical Technology
Background:
- Traditional razor blade slicing of brain tissue causes significant cellular damage.
- This damage creates diffusion barriers, delaying neuronal responses to drugs and hindering research.
Purpose of the Study:
- To introduce and evaluate a macromingotome for cutting brain slices using water jets.
- To assess the preservation of neuronal structure and function in slices prepared with this new method.
Main Methods:
- A macromingotome was developed to cut nervous tissue with high-pressure physiological saline jets.
- Hippocampal slices (300-500 microm) were cut using varying pressures (100-1800 bar) and nozzle diameters (35-100 microm).
- Optimal cutting parameters were identified as 400-600 bar with 60-80 microm nozzles.
Main Results:
- Intact CA1, CA3 neurons, and granule cells were observed less than 10 microm from the surface using infrared microscopy.
- Light microscopy confirmed intact fine structures in superficial neurons.
- Electrophysiological recordings showed normal membrane potentials, full action potentials, and stable epileptiform discharges.
Conclusions:
- The macromingotome provides a viable alternative to traditional slicing methods.
- This technique preserves superficial neuron viability and function, improving access for electrophysiological and neuropharmacological studies.
