The Subcutaneous Transmitter A3028, and its predecessors the A3019 and A3013, are wireless sensors designed to monitor biometric signals within live animals. We describe their operation in Subcutaneous Transmitter System.
Here is a list of available electrodes for use with our subcutaneous transmitters.
|A||Bare wire, length 2 mm, stainless steel helix.|
|B||Screw, thread 0-80, length 3.2 mm.|
|C||Screw, thread M0.5, length 0.6 mm.|
|D||Pin, diameter 0.30 mm, length 3.1 mm, Mill-Max 4353-0-00-15-00-00-33-0|
|E||Socket, for pin diameters 0.20-0.33 mm, Mill-Max 4428-0-43-15-04-14-10-0|
|F||Pin, diameter 0.64 mm, length 4.1 mm, Mill-Max 5035-0-00-15-00-00-33-0.
Mates with Plastics One socket E363/0.
|G||Pin, diameter 0.51 mm, length 6.0 mm, Mill-Max 5063-0-00-15-00-00-33-0.
Mates with Plastics One socket MS303/6.
|H||Depth electrode, wire 125-μm dia Pt-Ir, insulation 200-μm dia teflon.
Connect with socket, locate with guide cannula, remove after cementing.
|J||Depth electrode, wire 125-μm dia 316SS, insulation 200-μm dia teflon.
Connect with socket, locate with guide cannula, remove after cementing.
We discuss the encapsulation of implantable transmitters in Encapsulation. We discuss the design and manufacture of their silicone-insulated, helical, stainless-steel leads in Flexible Wires. The electrical contacts at the ends of these leads are what we call the electrodes. The primary purpose of the electrodes is to detect a biometric potential for transmission and recording. We designed the SCT system to monitor the electroencephalograph (EEG) or local field potential (LFP) in a live animal. For a discussion of the sources of the EEG and LPF signals, see The Source of EEG. Here we present the performance of various electrode in recording the extracellular potential within the cortex of a live animal.
[05-FEB-16] The flexible leads of a subcutaneous transmitter contain a stainless steel helix to carry biometric signals to the transmitter's amplifier. The wire itself is 316 stainless steel, which resists corrosion well, but will not accept solder unless treated with acid flux.
The bare-wire electrode is attractive because the experimenter can re-create the electrode without using solder or acid flux. We must take care to anchor the bare wire securely with respect to the recording point. It is not sufficient to put the wire into a skull hole and hold it in place with dental cement. The best way to hold the wire in place is with a screw in the same hole as the wire.
We start by stretching the exposed steel helix until it is nearly straight. We cut it to a length that will reach through a skull hole and penetrate the brain to the correct depth, or rest upon the surface of the brain. A penetration of 1 mm gives a more powerful EEG signal than a wire tip on the dura. In rats, we recommend a length of 5 mm. In mice, 2.5 mm. We bend the wire by ninety degrees half-way along its length. We place the wire in a skull hole and lay the insulated lead along the surface of the skull. We thread a screw into the hole. The exposed wire on top of the skull allows the screw to avoid interfering with the silicone insulation of the lead, and will allow dental cement to bond to the wire directly. In rats, the 2.5 mm wire passing through the skull will penetrate roughly 1 mm into the brain. In mice, the penetration of the 1.2-mm wire tip will be similar, because the skull is thinner.
With the wire held in place by a screw, we cover the screw with dental cement. The cement anchors and insulates the head of the screw and bonds to the wire itself. The helix will flex between the transmitter and the anchor screw, but the bare wire and the screw will be fixed with respect to the skull. Movement of the brain with respect to the skull will generate movement artifact, but we will not see sudden steps due to intermittent metal-on-metal contact. Nor will we see EMG artifact from muscles above the screw, because we have insulated the top of the screw with cement.
When we retrieve the transmitter from the animal, we tend to cut the leads where they emerge from the skull cement. It is possible to dissolve dental cement with vinegar, but usually the animal's brain is needed intact for examination. Having cut the leads, we remove silicone from the tips to expose more wire, and so we can re-create the bare wire electrode for a second implantation.
We have tried many other variations on the bare-wire electrode described above, and all of them exhibit movement artifact that is common enough to hinder seizure detection. For example, we tried stripping the insulation off 20-mm of wire and fastening this wire to the skull surface with two screws to act as a reference electrode. This arrangement generates transients. We clap our hands, the animal flinches, and the EEG jumps by 20 mV.
The bare wire is currently our most popular electrode for EEG recordings, because of the flexibility it offers. But if we want to look deeper into the brain, we cannot use the bare helix, because it has no insulation, and it is not straight enough to locate precisely.
[05-FEB-16] Stainless steel screws are useful as electrodes for EEG recording, because they are easy to anchor to the skull. We drill a hole slightly smaller than the screww, and thread the screw into the hole. The steel wire of the pick-up lead is soldered to the head of the screw. If the screw is large enough, we solder the wire to one side of the slot in the screw head. During implantation, we turn the screw with the slot. But of course we have a pick-up lead connected to the screw, so the lead rotates along with the screw. In order to leave the pick-up lead relaxed when the screw is in place, we must pre-twist it by the same number of turns required to thread the screw down into the hole. If the screw is small enough, we can simply press the threads into the skull hole, hold the screw in place with cement, and in the days following, the skull will grow into the threads of the screw and hold it firmly in place. Thus we avoid, with the smaller screw, any pre-twisting of the lead.
The screw is in electrical contact with the wire at its head, with the skull it passes through, and with the brain at its tip. We cover the head of the screw with dental cement, so it is insulated from body fluids and from any muscles that may lie over it, such as is the case for lateral skull screws in rats. Because the conductivity of the skull is low compared to that of the brain, the screw, despite its large and clumsy appearance, provides a connection to the surface of the brain. We have used skull screws in hundreds of implantations, and they provide reliable, artifact-free recordings. Despite the need to pre-twist the leads for larger screws, the skull screw remains the easiest electrode with which to obtain a reliable recording.
The following table summarizing the dimensions and properties of the screws we have used with our transmitters.
|Name||Diameter (mm)||Thread Pitch (mm)||Length (mm)||Applications|
|0-80 1/8"||1.52||0.32||3.2||Rat skull when >8 weeks.|
|00-90 1/16"||1.2||0.28||1.6||Rat skull when <8 weeks.|
|000-120 1/16"||0.86||0.21||1.6||Rat or mouse skull electrode.|
|M0.5-0.125-1.0||0.50||0.125||1.0||Mouse skull electrode.|
|M0.5-0.125-0.6||0.50||0.125||0.6||Mouse skull electrode.|
[04-AUG-10] We obtain samples of a screw, 00-96X1-16 from Plastics One, from CHB. With the help of acid flux, we solder screws to the tips of the wires of an A3013A, as shown here. The wires are soldered to the top sides of the heads of the screws. We tried soldering the wires to the underside of the screw heads, but we found that solder spread all down the length of the screw, obscuring the threads. Even when we solder the wire to the top of the head, we must take care not to allow solder to spread to the threads, and we do this by threading the screw into a small hole in a piece of box cardboard. The cardboard also provides us with a means of holding the screw while we make the joint.
[12-AUG-10] Archive M1281455686 contains EEG recorded by screws in No8 compared to EEG recorded by epoxy contacts in No10. The screws provide a stable baseline. Both contacts provide wave burst and seizure detection. The screws appear to be slighly less sensitive to seizure activity than silver epoxy or the bare wire electrodes. Peak seizure power in the 2−10 Hz range is around 5000 k counts-sq compared to 10,000 k counts-sq for the other electrodes.
[15-AUG-10] CHB implants No6 with screws. The wires of this transmitter are only 100 mm long, a bit short. We see large transient jumps on the signal. Sameer reports, "I think the unstable baseline from transmitter#6 can be attributed to the use of epoxy to secure one of the screws instead of a drilled hole. Due to some reason, I think size or location of leads, YingPeng couldn't drill it in. However, the other screw was fastened into the skull."
[25-AUG-10] We receive two sizes of 303 stainless steel screws from SmallParts (now Amazon Supply). Both have what is called the binding head, which is flatter than the screws we started with, as you can see in the photograph below.
All these screws are 1.6 mm long (1/16"). The 000-120 has diameter 0.86 mm with thread pitch 0.21 mm. The 00-90 has diameter 1.2 mm with thread pitch 0.28 mm. the 00-96 has diameter 1.2 mm and thread pitch 0.26 mm.
[03-APR-11] We have settled upon the 00-90, 1.6-mm long binding head screw as the standard for the A3019D transmitter for implanting in rats. For mice, Louise Upton has chosen an M0.5-0.125 screw, the B0038QOYFA, which has a thread 1.1 mm long and 0.5 mm in diameter, with pitch 0.125 mm. We soldered this screw to the 70-mm leads of an A3019A. She drilled holes in the skull of a dead mouse and declared the screw to be the perfect size. She did not thread the screw into the hole, but instead pushed it in. The threads held the screw in place.
[03-APR-11] We study 15 hours of recordings from ION/UCL, from eight control animals. The transmitters are A3019Ds with 00-90 binding head screws 1.6 mm long. The leads are insulated right up to the solder joints on the screw head. The screws are set directly into the skull with dental cement to hold them in place and insulate them. For a typical recording from eight sets of screw electrodes, see here.
We see few artifacts in the EEG. The amplitude of each trace is around 40 μV. Another similar period of recordings from eight different animals with the same electrodes shows 40 μV baseline EEG also. The consistency between the baseline amplitudes suggests that the screws are giving a repeatable contact with the brain.
[04-APR-11] ION/UCL suspects that the 00-90 screws are coming loose from the skull after a few weeks. "Yes, I also had the impression that the power per frequency band was quite consistent between animals, at least during the first week of recording. After that power tends to decrease across all bands in quite a lot of animals. This is probably caused by the skull getting thicker and slowly pushing the screws out. This ends in rats loosing their electrodes after 2 to 4 weeks. I've now soldered larger screws on a couple of transmitters and will also add a few more screws just to anchor the headpiece to see if this will enable recordings with stable band power for two or three months."
[13-SEP-12] By now we have supplied to ION over thirty transmitters with 00-80 screws, 1.6 mm in diameter and 3.2 mm long. These screws require a hole in the skull of diameter roughly 1.2 mm. They appear to work well for implants greater than four weeks, when the smaller screws lose sensitivity.
[12-JUN-13] Our M0.5-0.125-1.0 screw is $8 each in quantity 100, and it is often out of stock at the supplier. By now, Louise Upton and Sukhvir Wright of Oxford University have implanted roughly forty transmitters with these screws, and they found them to be too long. They use a 0.4-mm thick washer to back the screw away from the top of the skull. The bone is 0.4 mm thick, so the tip of the screw would be roughly 0.2 mm into the brain, pressing upon the surface. Today we ordered, from US Microscrew, quantity 1000 of a custom-made screw, diameter 0.5 mm, shaft length 0.6 mm, which we name M0.5-0.125-0.6.
[23-JUL-13] We receive our 1000 screws and find that their threads are fine, but the heads are mangled by the slot cut, as shown here (US Microscrew, 0.5 mm diameter, 0.6 mm length screw, first attempt).
[02-AUG-13] We receive from US Microscrew another 1000 screws to replace the first set. This lot don't have a slot cut in the head. We cover the entire head with solder, so we don't need the slot. Removing the slot makes the part easier to make.
[05-FEB-16] These M0.5 threaded electrodes have provided tens of thousands of hours of reliable recordings.
[05-FEB-16] Gold-plated pins allow us to connect the end of our EEG lead to depth electrodes. The depth electrode is too cumbersome to pass easily beneath the skin of an animal from the transmitter to the head. The pin, on the other hand, is easy to move up under the skin. Our contact pins are made by Mill-Max, and are summarized in their data sheet. The photograph below shows our D Electrode, which mates with our H and J Electrodes.
The pins are easy to solder to a steel wire. They provide a gold-plated finish, which mates well with the gold-plated finish of a socket, to provide a corrosion-resistant electrical contact within dental cement saturated with water vapor.
Our F and G eletrodes are pins that mate with electrodes made by Plastics One.
When you choose an electrode with a socket, we will look at the socket drawing and choose a pin that will fit sungly and take up as little vertical height as possible, to keep your implantation compact.
[15-JUN-16] To reach farther into brain, we need a stiff, insulated wire. A depth electrode must provide a means to hold the wire securely during implantation so we can move it to the correct depth. To confirm that the wire tip is in the correct location, we may need to examine the local field potential around the wire tip as we lower it into the brain. The photograph below shows our J-Electrode, which provides a stainless steel wire. Our H-Electrode provides a platinum-iridium wire.
The pick-up lead runs along the skull surface and attaches to the depth wire. We place an anchor screw nearby. The anchor screw and the depth electrode will be part of the same body of dental cement, so the anchor screw will secure the depth electrode. The socket is soldered to the electrode wire. For the H-Electrode's stainless steel wire, we use acid flux to make the joint. The socket has a hole at each end. The pin may be inserted only at one end. When, in our view, the socket is below the wire, the entry hole is the one facing us. When the socket is above the wire, as in the view below, the entry hole is facing away from us, and the tip of the pin will emerge from the hole we see.
The guide cannula permits the implanter to lower the electrode into the brain while monitoring the EEG signal. Once the electrode is in position, we cover the socket and anchor screw with dental cemant nd let it cure. We take care not to bury the tip of the guide cannula in the cement. Once the cement is cured, we cut the Pt-In wire where it emerges from the top of the guide cannula. We raise the guide cannula off the wire. We cut the wire where it emerges from the cement. We cover the exposed tip of the wire with more cement. Now the depth electrode is fastened to the anchor screw by dental cement, and there are no exposed metal contacts.
[08-MAR-16] Here are instructions for implanting a J- or H-Electrode. Our collaborators at ION/UCL took these instructions and composed these Pictoral Instructions, which you may find more useful than our written list.
|1||Remove from gel pack and inspect electrode. Locate the opening on the socket.|
|2||Press transmitter lead D-Pin into the socket. If small plug comes out the other side, discard.|
|3||Remove D-Pin from socket. Be careful not to bend the wire.|
|4||Cut wire to length. From socket to tip is skull thickness plus depth.|
|5||Sterlize with alcohol. If kapton tape comes off, leave it off.|
|6||Straighten wire if necessary. It may be bent at the end of the guide cannula.|
|7||Mount guide cannula thread in stereotactic clamp. Grip on the tape, if it is still attached.|
|8||Implant transmitter and insert D-Pin into socket. Be sure to insert into the correct end of the socket.|
|9||Lower wire into skull hole until socket rests upon skull. The electrode lead will be running along the skull surface.|
|10||Coat skull around socket with VetBond surface treatment. You can skip this step if you do not have VetBond.|
|11||Cover socket with dental cement. Do not allow the guide cannula to become fixed in the cement.|
|12||Wait for dental cement to cure, then cut wire loop at top of guide cannula.|
|13||Raise up the clamp to pull guide cannula off the wire, leaving wire sticking up.|
|14||Cut the wire flush with the top of the dental cement.|
|15||Cover the wire stub with dental cement.|
[26-SEP-15] At Edinburgh University, they implant the prototype H-Electrode in trangenic mice. The figure below shows depth and surface field potentials recorded simultaneously with a dual-channel subcutaneous transmitter. The surface and reference electrodes are 0.5-mm screws.
When implanting the H-Electrode, the Edinburgh group monitored EEG from the H-Electrode while lowering the tip of the electrode into the hippocampus. They adjusted the depth of the electrode until they obtained the largest possible EEG amplitude. The 125-μm tip of the depth electrode has impedance roughly 10 kΩ, compared to 2 kΩ for the screw. The depth recording has baseline amplitude 400 μV compared to 80 μV for the surface recording. There we see one or two step artifacts per hour in these dual-channel recordings. But the artifacts are shared between the depth and surface recordings, like this, so they cannot be the result of movement of the H-Electrode alone.
[08-AUG-16] We receive archive M1470389836.ndf from ION/UCL. Two A3028H-DDC transmitters are implanted in two conscious, freely-moving mice with two J-Electrodes each. Amplitude is roughly 80 μV from all channels. We look through the hour-long recording and see none of the step artifacts that motivated the development of the H and J-Electrodes.
We note that ION/UCL did not adjust the depth of the J-Electrode to obtain the maximum amplitude of EEG, but rather set the depth to a certain value during implantation without monitoring EEG.
[05-FEB-16] Conductive epoxy might allow us to cement a bare wire directly onto the skull in two places so as to provide EEG recording without drilling any holes. We tried silver epoxy, as we describe below. The recordings pick up muscular activity from the scalp of the host rat, which we see also in un-insulated screw electrodes. Such activity we can detect and classify in order to distinguish it from EEG. The recordings showed continuous drift in the baseline voltage, not by transient steps as we see in poorly-secured wire electrodes, but as a drift of several millivolts over one-second intervals. Such drifts we can overcome with our event classification. The conductive epoxy recordings are, we believe, adequate for event detection. But the electrodes themselves are not as reliable as screws, in that they tend to come loose. More work with silver epoxy might increase reliability, but such work has not yet occurred.
[12-AUG-10] CHB implanted transmitter No10 and glued the ends of the analog wires to the animal's skull using silver epoxy, which provides an electrically conducting contact. With these contacts, we observed seizure-like activity in a non-epileptic rat, as shown here in archive M1281121460. We are convinced that the epoxy contacts are picking up EEG. Later, in archive M1281455686, we can compare No10's contacts with No8's steel screw contacts. When we say baseline signal we mean the signal low-pass filtered to 1 Hz or so, which is a measure of the average value of the signal over a fraction of a second.
From this one experiment, we see the baseline swinging up and down compared to the stable baseline we obtain from the screws. But the silver epoxy contact sensitivity compares well to that of the bare wires, and exceeds that of the screw contacts.
The following plot shows power in the 2−20 Hz band for the two transmitters during three hours of recording.
The peak power from No10 corresponds to vigorous seizures like this one, not any transient spikes or level shifts. In addition to seizure detection, the silver epoxy contacts are also sensitive to wave bursts in the 40−160 Hz band. The plot below shows simultaneous wave-bursts from No8 (screws) and No10 (epoxy).
It appears that the silver epoxy has only one disadvantage compared to screws: a moving baseline. But the moving baseline does not appear to interfere with seizure or wave burst detection, and the silver epoxy method does not require holes in the skull.
[30-MAY-17] As we mention several times in the discussion above: the movement of electrodes with respect to the surrounding animal tissue will add step-changes in potential to the EEG signal. These step changes pass through the transmitter's high-pass filter and come out looking like pulses roughly half a second long. The theoretical step response of the A3028B, for example, is the blue graph shown here. If we solder the two leads of a transmitter together, drop the transmitter in water, and stir it around, he standard deviation of the signal is less than 20 μV. We see no step artifact. If we separate the leads and allow them to measure the variation in electric potential in the water as we stir, we see 1-mV rumble, but not sharp steps. It is when we allow the eletrodes to make intermittent contact with one another as we stir the tranmsitter in water that we start to see 10-mV steps being added to the signal. We call these steps "movement artifact", and they are due to changes in the electrode potential. Changes in the chemistry of the conducting solutions between the metal electrodes can generate step changes of over one hundred millivolts.
Movement artifact arises in EEG recordings when the electrodes are not secured with respect to the skull. As implanters have learned to secure wires and screws more effectively to the animal skull, the frequency of these artifacts has decreased. In mice, these artifacts are particularly difficult to eliminate. Here are some examples of movement artifact from mouse recordings.
Here we see pulses caused by steps of order 10 mV, which are shaped into pulses by the transmitter's high-pass filter. In the figure below we see far larger artifact from the same M0.5 skull screws in another mouse.
Here we see large artifact with more complex progression, also from skull screws in a mouse. The amplitude is 15 mVpp.
We are not certain of the origin of the following features. The two traces are from a dual-channel transmitter implanted in a mouse. If the electrode potential of the common electrode changes, we expect the effect upon the X and Y channels to be shared. If the electrode potential of either X or Y changes, we expect to see the effect only on X or Y. Here we see opposite changes in X and Y.
The number of movement artifacts in a recording varies greatly from one implant to the next, and is a strong function of who performs the surgery. Recordings from rats tend to have fewer than one artifact per hour for all experienced implanters. Movement artifact in mice varies from fewer than one per hour to hundreds of artifacts per hour. Those implanters whose recordings are almost free of artifact have reduced the artifact rate by working on better anchoring of their electrodes on the mouse skull. We are currently working on developing a guide to successful implantation in mice.
Our various users have been able to make every type of electrode termination work well, with the exception of the silver epoxy. In the A3028 manual page, you will find a table giving letter codes for the types of bare wire, screw, and pin in current use with our customers.