Electrode Fabrication and Implantation in Aplysia californica for Multi-channel Neural and Muscular Recordings in Intact, Freely Behaving Animals

Summary

A technique is described for implanting four in vivo electrodes to monitor the neuromuscular control of feeding behavior in Aplysia californica.

Abstract

Recording from key nerves and muscles of Aplysia during feeding behavior allows us to study the patterns of neural control in an intact animal. Simultaneously recording from multiple nerves and muscles gives us precise information about the timing of neural activity. Previous recording methods have worked for two electrodes, but the study of additional nerves or muscles required combining and averaging the recordings of multiple animals, which made it difficult to determine fine details of timing and phasing, because of variability from response to response, and from animal to animal. Implanting four individual electrodes has a very low success rate due to the formation of adhesions that prevent animals from performing normal feeding movements. We developed a new method of electrode fabrication that reduces the bulk of the electrodes inside the animal allowing for normal feeding movements. Using a combination of glues to attach the electrodes results in a more reliable insulation of the electrode which lasts longer, making it possible to record for periods as long as a week. The fabrication technique that we describe could be extended to incorporate several additional electrodes, and would be applicable to vertebrate animals.

Protocol

1. Electrode Fabrication

1. To create the electrode, use scissors to cut a piece of enamel-coated 0.001 inch diameter stainless steel wire about 2 feet long. Attach a small ball of putty to each end of the wire, fold the wire in half, and spin the ends to create a twisted pair, called a differential electrode.
2. Tape both ends of the wire to prevent unraveling. Make four differential electrodes, using a different color of tape for each differential electrode for identification purposes (Figure 1A).
3. Twist two of the differential electrodes together, leaving two inches untwisted on one end and 1 inch on the other end. Repeat with the remaini....

Discussion

A key innovation that has made these recordings successful is combining the electrodes into one cable. Two electrode recordings have been used before, but expanding to four individual electrodes resulted in the formation of surgical adhesions that often prevented the animal from making normal feeding movements. Combining the electrodes into one cable reduces the bulk of the electrodes inside the animal, which in turn reduces the immune response and allows the animal to move more freely.

The o.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Enamel coated stainless steel wire	California Fine Wire Company		0.001D, coating h
Household Silicone II Glue	GE Healthcare
Kwik-Sil Adhesive	World Precision Instruments, Inc.
Duro Qwik-Gel superglue	Henkel Corp
A-M Systems model 1700 amplifier	A-M Systems		Filter settings:100-1000Hz nerves,10-1000Hz I2 muscle
Axoscope 10.1	Molecular Devices
Gold Connector Pins	Bulgin	SA3148/1
Gold Connector Sockets	Bulgin	SA3149/1