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A simple surgical procedure for immobilization of rodent eyes
TECHNICAL
NOTE
A SIMPLE SURGICAL PROCEDURE FOR IMMOBILIZATION OF RODENT EYES R. B. H.
TOOTELL
and
GERALD
H.
JACOBS
Department of Psychology, University of California. Santa Barbara. California 93106. U.S.A. (Receiced
27 April
INTRODUCI’ION
Because previous methods for reducing the occurin rodents during recording experiments have some drawbacks, we have used a surgical procedure to interrupt the oculomotor input to the eye at the level of the entry of these nerves into the orbit. This takes only a few minutes and it restricts eye movements to a level that is roughly commensurate with that achieved in paralyzed preparations (Cicerone and Green. 1977). The technique has been used successfully in a large number of ground squirrels and rats, and could probably be adapted for use with other rodent species. The procedure is outlined here for the rat.
rence of the eye movements
PROCEDURE
The rat is anesthetized and placed in a stereotaxic head holder. The eyelids are held open with single sutures. The temporal canthus is cut back about 2-3 mm and then pierced with closed iris forceps at a location immediately posterior to the fasci bulbi. The foraps are inserted through the orbital fat until the tips reach the posterior orbit where they are opened to widen the hole. The IIIrd, IVth and VIth nerves enter the orbit in a common bundle through the anterior lacerated foramen (ALF). The ALF is very slightly dorsal to the zygomatic arch. A lesioning electrode pushed horizontally through the entry hole, hugging the posterior orbit and the zygomatic arch. will usually come to rest in the immediate vicinity of the ALF. In the rat, the posterior limit of the ALF is a bony ridge which is also slightly ventral, forming a landmark. When positioning the electrode is is important to keep the electrode tip held resolutely horizontal, or even slightly ventrally. This procedure helps to keep the electrode away from the optic foramen which is located on the roof of the medial orbit. Figure 1 illustrates the location of the electrode in the orbit when it is appropriately-positioned for lesioning. The lesioning electrode was ma& from a 17 guage stainless steel cannula. The tip of this cannula was bent at an angle of about 70” and then crushed flat. The tip was rounded ofi smoothly. The electrode was insulated with formvar except for an area of 2-3 mm* on the bottom of the electrode tip. Since the electrode is constructed to conform to the shape of the ALF, separate electrodes are required for left and right eyes. After the electrode tip had been lodged in the dorsal ALF using the approach described above, the electrode was connected to the negative pole of a IO V d.c. supply and the circuit was completed via a screw embedded in the skull of the animal. Lesions were made by passing 30 rn~
1979)
of current for 3-5 sec. To insure a complete destruction of the oculomotor nerves. several such lesions are usually made with the lesioning electrode successively moved to slightly different positions. RESULTS
The surgical procedure appears to produce no deleterious effects on the retina since the electroretinogram recorded from lesioned animals did not differ from that of control animals and since a histological examination of the retina revealed no obvious signs of damage. The degree of immobilization
produced by the surgical technique was assessed by measuring eye movements several days following the surgery. The animal was anesthetized with urethane to a level where the unstimulated animal would remain quiescent, but still give vigorous pawing movements to a mild ear pinch. This level was generally achieved in adult male rats with a dose of about 1.8 g/kg. For recording, the head
Fig. I. Dorsal view of the rat skull with the cranial cavity exposed. The proper position of the lesioning electrode is illustrated in the right orbit. (OF: optic foramen: Zyg.: zygomatic arch; ALF. anterior lacerated foramen).
1281
B
I
I
Fig. 2. Plots of eye movements recorded from rodents following surgical immobilization of the eye. The dots i&icate eye position at successive IO min intervals. Traces (A) and (D) were obtained from rats 3 days after the lesioning procedure. Trace (B) is from a California ground squirrel 3 days following the lesion. Trace (C) was obtained from a rat l-3 hr following the surgery. See text for further discussion.
was painlessly immobilized in a fixed position by attaching some previously implanted skull screws to a rigid metal arm. A small mirror chip, 1.3 mg in weight, was attached to the cornea with Histoacryl tissue adhesive (TriHawk International. Montreal, Canada). A laser beam was reflected from this chip onto a tangent screen positioned 40 cm from the eye. The position of the reflected spot was tracked, typically with sampling intervals of 5 or 10 min. Figure 2 (A and D) shows records of eye movements obtained from urethane-anesthetized rats whose eyes had been surgically immobilized. Over the recording periods, which ranged from 1 to 3 or more hours, the region over which the eye moved did not exceed I’. This value can be compared to average excursions of 4’ or greater over 10 min time periods for other rats identically tested but without prior surgical immobilization. The results shown in Fig. 2 are quite typical of those obtained from a number of other animals. Figure 2B also shows the residual eye movements recorded from a urethane-anesthetized ground squirrel after the surgical procedure. As for the rats, eye movements in ground squirrels are restricted to regions of less than l2 following the operation. In this case the degree of restriction of eye
mobement achlebed jurg!<:t:!;, iI :i;n gr<:it or more days between the time of the Iesioning and the recording date. If eye movements are compared in dnimals immediately following the lesion with those seen in animals examined 3 days or more after the lesion. it is found that although movements over an hour or more were not more than 1’ in both cases. those Jnimals which have just been lesioned often show long-term slow drifts which move thr eyes over somewhat greater excursions (see Fig. 3Ci. This problem seems to be more acute in ground squirrs!j than in rats. In any case. a three day wait has proved to be sufficient to eliminate the draft problem. In several cases single-cell recordings have been made from painlessly immobilized. unanesthetized ground squirrels whose oculomotor inputs had been previously lesioned. In these instances the degree of immobilization was sometimes poorer than in similarly-treated anesthetized animals. although still constderably superior to anesthetized animals which had not received lesions. The main problem in unanesthe-
tized (and under-anesthetized) animals is that the mandible of the rodents invades the ventral portion of the orbit so that any jaw movements are translated directly into eye movements. been noted in unanesthetized
A similar problem
has
primates with surgically immobilized eyes (Snodderly t’r al.. !978).
We have found in the course of doing recording experiments on anesthetized rats and ground squirrels that eye movements can be substantially curtailed through a simple surgical procedure which involves lesioning the oculomotor nerves at the point where they enter the orbit. Although the degree of immobilization achieved in this way is perhaps slightly less than that obtained using neuromuscular blocking agents, the simplicity of the procedure. in conjunction with the large receptive fields typically found in rodent visual systems, suggests this technique may prove more than adequate for many different purposes. Ackno&dgement-This work was supported by a grant from the National Eye Institute (EY-00105). REFEREXCES
Cicerone C. M. and Green D. G. (1977) Control of eye movements while recording from single units in the pigmented rat. Vision Res. 17. 985-987. Snodderly D. M.. Swadlow H. A. and Barlow R. B. (1978) Evaluation of a surgical method for immobilizing the eye of an alert monkey. Expl Brain Res. 31. 179-191.
NOTE
A SIMPLE SURGICAL PROCEDURE FOR IMMOBILIZATION OF RODENT EYES R. B. H.
TOOTELL
and
GERALD
H.
JACOBS
Department of Psychology, University of California. Santa Barbara. California 93106. U.S.A. (Receiced
27 April
INTRODUCI’ION
Because previous methods for reducing the occurin rodents during recording experiments have some drawbacks, we have used a surgical procedure to interrupt the oculomotor input to the eye at the level of the entry of these nerves into the orbit. This takes only a few minutes and it restricts eye movements to a level that is roughly commensurate with that achieved in paralyzed preparations (Cicerone and Green. 1977). The technique has been used successfully in a large number of ground squirrels and rats, and could probably be adapted for use with other rodent species. The procedure is outlined here for the rat.
rence of the eye movements
PROCEDURE
The rat is anesthetized and placed in a stereotaxic head holder. The eyelids are held open with single sutures. The temporal canthus is cut back about 2-3 mm and then pierced with closed iris forceps at a location immediately posterior to the fasci bulbi. The foraps are inserted through the orbital fat until the tips reach the posterior orbit where they are opened to widen the hole. The IIIrd, IVth and VIth nerves enter the orbit in a common bundle through the anterior lacerated foramen (ALF). The ALF is very slightly dorsal to the zygomatic arch. A lesioning electrode pushed horizontally through the entry hole, hugging the posterior orbit and the zygomatic arch. will usually come to rest in the immediate vicinity of the ALF. In the rat, the posterior limit of the ALF is a bony ridge which is also slightly ventral, forming a landmark. When positioning the electrode is is important to keep the electrode tip held resolutely horizontal, or even slightly ventrally. This procedure helps to keep the electrode away from the optic foramen which is located on the roof of the medial orbit. Figure 1 illustrates the location of the electrode in the orbit when it is appropriately-positioned for lesioning. The lesioning electrode was ma& from a 17 guage stainless steel cannula. The tip of this cannula was bent at an angle of about 70” and then crushed flat. The tip was rounded ofi smoothly. The electrode was insulated with formvar except for an area of 2-3 mm* on the bottom of the electrode tip. Since the electrode is constructed to conform to the shape of the ALF, separate electrodes are required for left and right eyes. After the electrode tip had been lodged in the dorsal ALF using the approach described above, the electrode was connected to the negative pole of a IO V d.c. supply and the circuit was completed via a screw embedded in the skull of the animal. Lesions were made by passing 30 rn~
1979)
of current for 3-5 sec. To insure a complete destruction of the oculomotor nerves. several such lesions are usually made with the lesioning electrode successively moved to slightly different positions. RESULTS
The surgical procedure appears to produce no deleterious effects on the retina since the electroretinogram recorded from lesioned animals did not differ from that of control animals and since a histological examination of the retina revealed no obvious signs of damage. The degree of immobilization
produced by the surgical technique was assessed by measuring eye movements several days following the surgery. The animal was anesthetized with urethane to a level where the unstimulated animal would remain quiescent, but still give vigorous pawing movements to a mild ear pinch. This level was generally achieved in adult male rats with a dose of about 1.8 g/kg. For recording, the head
Fig. I. Dorsal view of the rat skull with the cranial cavity exposed. The proper position of the lesioning electrode is illustrated in the right orbit. (OF: optic foramen: Zyg.: zygomatic arch; ALF. anterior lacerated foramen).
1281
B
I
I
Fig. 2. Plots of eye movements recorded from rodents following surgical immobilization of the eye. The dots i&icate eye position at successive IO min intervals. Traces (A) and (D) were obtained from rats 3 days after the lesioning procedure. Trace (B) is from a California ground squirrel 3 days following the lesion. Trace (C) was obtained from a rat l-3 hr following the surgery. See text for further discussion.
was painlessly immobilized in a fixed position by attaching some previously implanted skull screws to a rigid metal arm. A small mirror chip, 1.3 mg in weight, was attached to the cornea with Histoacryl tissue adhesive (TriHawk International. Montreal, Canada). A laser beam was reflected from this chip onto a tangent screen positioned 40 cm from the eye. The position of the reflected spot was tracked, typically with sampling intervals of 5 or 10 min. Figure 2 (A and D) shows records of eye movements obtained from urethane-anesthetized rats whose eyes had been surgically immobilized. Over the recording periods, which ranged from 1 to 3 or more hours, the region over which the eye moved did not exceed I’. This value can be compared to average excursions of 4’ or greater over 10 min time periods for other rats identically tested but without prior surgical immobilization. The results shown in Fig. 2 are quite typical of those obtained from a number of other animals. Figure 2B also shows the residual eye movements recorded from a urethane-anesthetized ground squirrel after the surgical procedure. As for the rats, eye movements in ground squirrels are restricted to regions of less than l2 following the operation. In this case the degree of restriction of eye
mobement achlebed jurg!<:t:!;, iI :i;n gr<:it
tized (and under-anesthetized) animals is that the mandible of the rodents invades the ventral portion of the orbit so that any jaw movements are translated directly into eye movements. been noted in unanesthetized
A similar problem
has
primates with surgically immobilized eyes (Snodderly t’r al.. !978).
We have found in the course of doing recording experiments on anesthetized rats and ground squirrels that eye movements can be substantially curtailed through a simple surgical procedure which involves lesioning the oculomotor nerves at the point where they enter the orbit. Although the degree of immobilization achieved in this way is perhaps slightly less than that obtained using neuromuscular blocking agents, the simplicity of the procedure. in conjunction with the large receptive fields typically found in rodent visual systems, suggests this technique may prove more than adequate for many different purposes. Ackno&dgement-This work was supported by a grant from the National Eye Institute (EY-00105). REFEREXCES
Cicerone C. M. and Green D. G. (1977) Control of eye movements while recording from single units in the pigmented rat. Vision Res. 17. 985-987. Snodderly D. M.. Swadlow H. A. and Barlow R. B. (1978) Evaluation of a surgical method for immobilizing the eye of an alert monkey. Expl Brain Res. 31. 179-191.