Stereotaxic surgery is a powerful technique utilized to manipulate the brain in living animals. Targeting regions deep within the intact brain poses a unique problem in that usually the surgical target cannot be located visually. Stereotaxy or stereotactic surgery was devised to target discrete regions of the brain by knowing their spatial relationships to visible landmarks. Using a three-dimensional coordinate system, tools can be directed to specific locations in order to measure brain activity, make a lesion, or perform genetic manipulations. This video will cover the principles of stereotaxic surgery, demonstrate essential steps of the procedure, and discuss various applications of the technique.
Before discussing the procedural details, let’s review the basic principles and tools of the technique. To start, stereotaxy depends on locating the targeted region of the brain on a three dimensional axis. For this purpose, a stereotactic atlas is used. This document is a 3D reconstruction of the brain obtained from serial sections of stained brains.
In this reference frame, spatial relationships are expressed using a set of 3 axes: Anterior-posterior, medial-lateral and dorsal-ventral. To determine the location of a region of interest, find the appropriate brain region in the atlas and use the grid to calculate the corresponding mediolateral and dorsoventral coordinates. The anteroposterior coordinate is obtained at the bottom or top of the page. This coordinate is measured from landmarks on the skull formed by the intersections of bone plates, which are known as sutures. The most prominent landmark, bregma, is defined as the point of intersection of the sagittal suture with the coronal suture. The point known as lambda is more posterior; where the sagittal suture intersects the lambdoid suture.
For accurate targeting to the calculated location during surgery, a device known as the stereotaxic frame is used . The frame consists of the main frame, the micromanipulator, and the probe holder, which is attached to the mounting clamp. Incisor and ear bars make contact with the animal in order to hold the head in a fixed position. The micromanipulator is used for precise movement of surgical probes to predetermined coordinates within the brain, and is equipped with sliding Vernier scales for exact distance measurements in all three dimensions.
To read a Vernier scale, first identify the location of the 0 line in relation to the tick marks on the right side of the scale. This side of the scale is labeled in 10 millimeter increments, so that 1 is 10 and 2 is 20. The location of the 0 mark here shows that the reading is in between 10 and 11 mm. To determine the first number to the right of the decimal point, find the line on the left scale that lines up best with a mark on the right side. Use the left scale to identify the value of this mark, which in this case is 9, making the final reading 10.9 mm.
You’re now familiar with the basic principles of stereotaxy, so let’s review how the surgical procedure is carried out. Before surgery, the scalp of an anesthetized animal is shaved and disinfected. To hold the animal in place on the stereotaxic frame, the ear bars are gently placed in the ears, and the front teeth are placed in the incisor bars. Eye ointment is applied to prevent corneal drying during surgery.
To expose the skull, make a small incision with a scalpel. Separate the muscle tissue gently and clean the surface of the skull. Then, use the micromanipulator to lower a probe to bregma and take note of its dorsoventral coordinate. Then raise the probe and repeat this procedure at lambda. If the lambda measurement is less than 0.1 mm from the dorsoventral coordinate at bregma, the skull is considered level and you are ready to proceed. If not, raise or lower the incisor bar and repeat the measurements.
Now that the brain is level, return to bregma and record the medio-lateral and antero-posterior coordinates. Based on the target coordinates from the brain atlas, calculate the movements required to reach that site and shift the probe to the target position. Mark this position on the skull and use a drill to carefully make a hole in the bone.
Just under the skull you will find the dura, a thick durable membrane that can be gently sectioned with the tip of a small needle.
Next, the micromanipulator arm is placed into position to lower the probe to the targeted dorsoventral coordinate, at which point the desired manipulation can be performed.
Now that you understand the basic steps of the procedure, let’s take a look at some of the many ways neuroscientists use stereotaxic surgery to answer questions in the lab.
To begin, stereotaxy is extremely useful for delivering experimental agents into precise brain regions. These agents may include tracer dyes used to visualize neuronal projections and connectivity, chemical compounds, or reagents for genetic manipulation of neurons. For instance, the injection of genetically engineered viruses will result in delivery of a gene of interest to infected cells. Here, a virus was injected into the hippocampus, resulting in GFP expression in a small population of neurons.
On the other hand, viral delivery of small interfering RNA can prevent the expression of desired proteins. In this example, expression of the protein SOD1 was suppressed in different brain regions following stereotaxic injection into the lateral ventricle.
Alternatively, stereotaxic surgery can be used to place electrodes in the brain for recording of the electrical signals that accompany neural activity. After the animal recovers from the surgical procedure, the electrodes can be connected to electrophysiological recording systems to observe neuronal firing while the mouse is freely behaving.
In addition to measuring neuronal activity, stereotaxic surgery is instrumental to microdialysis. This technique allows continuous monitoring of drug, neurotransmitter or metabolite concentrations in the brain tissue of awake animals. To do this, a small, fluid-filled probe with a semipermeable membrane at its tip is implanted into a specific brain region. Small solutes including neurotransmitters, hormones, and drugs diffuse into the probe, where they can be collected for analysis. In this experiment, glucose concentration in the hippocampus was measured as a readout for local neural activity while rats completed a task requiring the use of spatial working memory.
You’ve just watched JoVE’s video on stereotaxic surgery. In this video, we’ve covered the principles and methods of the procedure as well as its many applications in neuroscience labs.
Thanks for watching!
Stereotaxic (or stereotactic) surgery is a method used to manipulate the brain of living animals. This technique allows researchers to accurately targ…
Stereotaxic surgery is a powerful technique utilized to manipulate the brain in living animals. Targeting regions deep within the intact brain poses a unique problem in that usually the surgical target cannot be located visually. Stereotaxy or stereotactic surgery was devised to target discrete regions of the brain by knowing their spatial relationships to visible landmarks. Using a three-dimensional coordinate system, tools can be directed to specific locations in order to measure brain activity, make a lesion, or perform genetic manipulations. This video will cover the principles of stereotaxic surgery, demonstrate essential steps of the procedure, and discuss various applications of the technique.
Before discussing the procedural details, let’s review the basic principles and tools of the technique. To start, stereotaxy depends on locating the targeted region of the brain on a three dimensional axis. For this purpose, a stereotactic atlas is used. This document is a 3D reconstruction of the brain obtained from serial sections of stained brains.
In this reference frame, spatial relationships are expressed using a set of 3 axes: Anterior-posterior, medial-lateral and dorsal-ventral. To determine the location of a region of interest, find the appropriate brain region in the atlas and use the grid to calculate the corresponding mediolateral and dorsoventral coordinates. The anteroposterior coordinate is obtained at the bottom or top of the page. This coordinate is measured from landmarks on the skull formed by the intersections of bone plates, which are known as sutures. The most prominent landmark, bregma, is defined as the point of intersection of the sagittal suture with the coronal suture. The point known as lambda is more posterior; where the sagittal suture intersects the lambdoid suture.
For accurate targeting to the calculated location during surgery, a device known as the stereotaxic frame is used . The frame consists of the main frame, the micromanipulator, and the probe holder, which is attached to the mounting clamp. Incisor and ear bars make contact with the animal in order to hold the head in a fixed position. The micromanipulator is used for precise movement of surgical probes to predetermined coordinates within the brain, and is equipped with sliding Vernier scales for exact distance measurements in all three dimensions.
To read a Vernier scale, first identify the location of the 0 line in relation to the tick marks on the right side of the scale. This side of the scale is labeled in 10 millimeter increments, so that 1 is 10 and 2 is 20. The location of the 0 mark here shows that the reading is in between 10 and 11 mm. To determine the first number to the right of the decimal point, find the line on the left scale that lines up best with a mark on the right side. Use the left scale to identify the value of this mark, which in this case is 9, making the final reading 10.9 mm.
You’re now familiar with the basic principles of stereotaxy, so let’s review how the surgical procedure is carried out. Before surgery, the scalp of an anesthetized animal is shaved and disinfected. To hold the animal in place on the stereotaxic frame, the ear bars are gently placed in the ears, and the front teeth are placed in the incisor bars. Eye ointment is applied to prevent corneal drying during surgery.
To expose the skull, make a small incision with a scalpel. Separate the muscle tissue gently and clean the surface of the skull. Then, use the micromanipulator to lower a probe to bregma and take note of its dorsoventral coordinate. Then raise the probe and repeat this procedure at lambda. If the lambda measurement is less than 0.1 mm from the dorsoventral coordinate at bregma, the skull is considered level and you are ready to proceed. If not, raise or lower the incisor bar and repeat the measurements.
Now that the brain is level, return to bregma and record the medio-lateral and antero-posterior coordinates. Based on the target coordinates from the brain atlas, calculate the movements required to reach that site and shift the probe to the target position. Mark this position on the skull and use a drill to carefully make a hole in the bone.
Just under the skull you will find the dura, a thick durable membrane that can be gently sectioned with the tip of a small needle.
Next, the micromanipulator arm is placed into position to lower the probe to the targeted dorsoventral coordinate, at which point the desired manipulation can be performed.
Now that you understand the basic steps of the procedure, let’s take a look at some of the many ways neuroscientists use stereotaxic surgery to answer questions in the lab.
To begin, stereotaxy is extremely useful for delivering experimental agents into precise brain regions. These agents may include tracer dyes used to visualize neuronal projections and connectivity, chemical compounds, or reagents for genetic manipulation of neurons. For instance, the injection of genetically engineered viruses will result in delivery of a gene of interest to infected cells. Here, a virus was injected into the hippocampus, resulting in GFP expression in a small population of neurons.
On the other hand, viral delivery of small interfering RNA can prevent the expression of desired proteins. In this example, expression of the protein SOD1 was suppressed in different brain regions following stereotaxic injection into the lateral ventricle.
Alternatively, stereotaxic surgery can be used to place electrodes in the brain for recording of the electrical signals that accompany neural activity. After the animal recovers from the surgical procedure, the electrodes can be connected to electrophysiological recording systems to observe neuronal firing while the mouse is freely behaving.
In addition to measuring neuronal activity, stereotaxic surgery is instrumental to microdialysis. This technique allows continuous monitoring of drug, neurotransmitter or metabolite concentrations in the brain tissue of awake animals. To do this, a small, fluid-filled probe with a semipermeable membrane at its tip is implanted into a specific brain region. Small solutes including neurotransmitters, hormones, and drugs diffuse into the probe, where they can be collected for analysis. In this experiment, glucose concentration in the hippocampus was measured as a readout for local neural activity while rats completed a task requiring the use of spatial working memory.
You’ve just watched JoVE’s video on stereotaxic surgery. In this video, we’ve covered the principles and methods of the procedure as well as its many applications in neuroscience labs.
Thanks for watching!
Stereotaxic surgery is a powerful technique utilized to manipulate the brain in living animals. Targeting regions deep within the intact brain poses a unique problem in that usually the surgical target cannot be located visually. Stereotaxy or stereotactic surgery was devised to target discrete regions of the brain by knowing their spatial relationships to visible landmarks. Using a three-dimensional coordinate system, tools can be directed to specific locations in order to measure brain activity, make a lesion, or perform genetic manipulations. This video will cover the principles of stereotaxic surgery, demonstrate essential steps of the procedure, and discuss various applications of the technique.
Before discussing the procedural details, let’s review the basic principles and tools of the technique. To start, stereotaxy depends on locating the targeted region of the brain on a three dimensional axis. For this purpose, a stereotactic atlas is used. This document is a 3D reconstruction of the brain obtained from serial sections of stained brains.
In this reference frame, spatial relationships are expressed using a set of 3 axes: Anterior-posterior, medial-lateral and dorsal-ventral. To determine the location of a region of interest, find the appropriate brain region in the atlas and use the grid to calculate the corresponding mediolateral and dorsoventral coordinates. The anteroposterior coordinate is obtained at the bottom or top of the page. This coordinate is measured from landmarks on the skull formed by the intersections of bone plates, which are known as sutures. The most prominent landmark, bregma, is defined as the point of intersection of the sagittal suture with the coronal suture. The point known as lambda is more posterior; where the sagittal suture intersects the lambdoid suture.
For accurate targeting to the calculated location during surgery, a device known as the stereotaxic frame is used . The frame consists of the main frame, the micromanipulator, and the probe holder, which is attached to the mounting clamp. Incisor and ear bars make contact with the animal in order to hold the head in a fixed position. The micromanipulator is used for precise movement of surgical probes to predetermined coordinates within the brain, and is equipped with sliding Vernier scales for exact distance measurements in all three dimensions.
To read a Vernier scale, first identify the location of the 0 line in relation to the tick marks on the right side of the scale. This side of the scale is labeled in 10 millimeter increments, so that 1 is 10 and 2 is 20. The location of the 0 mark here shows that the reading is in between 10 and 11 mm. To determine the first number to the right of the decimal point, find the line on the left scale that lines up best with a mark on the right side. Use the left scale to identify the value of this mark, which in this case is 9, making the final reading 10.9 mm.
You’re now familiar with the basic principles of stereotaxy, so let’s review how the surgical procedure is carried out. Before surgery, the scalp of an anesthetized animal is shaved and disinfected. To hold the animal in place on the stereotaxic frame, the ear bars are gently placed in the ears, and the front teeth are placed in the incisor bars. Eye ointment is applied to prevent corneal drying during surgery.
To expose the skull, make a small incision with a scalpel. Separate the muscle tissue gently and clean the surface of the skull. Then, use the micromanipulator to lower a probe to bregma and take note of its dorsoventral coordinate. Then raise the probe and repeat this procedure at lambda. If the lambda measurement is less than 0.1 mm from the dorsoventral coordinate at bregma, the skull is considered level and you are ready to proceed. If not, raise or lower the incisor bar and repeat the measurements.
Now that the brain is level, return to bregma and record the medio-lateral and antero-posterior coordinates. Based on the target coordinates from the brain atlas, calculate the movements required to reach that site and shift the probe to the target position. Mark this position on the skull and use a drill to carefully make a hole in the bone.
Just under the skull you will find the dura, a thick durable membrane that can be gently sectioned with the tip of a small needle.
Next, the micromanipulator arm is placed into position to lower the probe to the targeted dorsoventral coordinate, at which point the desired manipulation can be performed.
Now that you understand the basic steps of the procedure, let’s take a look at some of the many ways neuroscientists use stereotaxic surgery to answer questions in the lab.
To begin, stereotaxy is extremely useful for delivering experimental agents into precise brain regions. These agents may include tracer dyes used to visualize neuronal projections and connectivity, chemical compounds, or reagents for genetic manipulation of neurons. For instance, the injection of genetically engineered viruses will result in delivery of a gene of interest to infected cells. Here, a virus was injected into the hippocampus, resulting in GFP expression in a small population of neurons.
On the other hand, viral delivery of small interfering RNA can prevent the expression of desired proteins. In this example, expression of the protein SOD1 was suppressed in different brain regions following stereotaxic injection into the lateral ventricle.
Alternatively, stereotaxic surgery can be used to place electrodes in the brain for recording of the electrical signals that accompany neural activity. After the animal recovers from the surgical procedure, the electrodes can be connected to electrophysiological recording systems to observe neuronal firing while the mouse is freely behaving.
In addition to measuring neuronal activity, stereotaxic surgery is instrumental to microdialysis. This technique allows continuous monitoring of drug, neurotransmitter or metabolite concentrations in the brain tissue of awake animals. To do this, a small, fluid-filled probe with a semipermeable membrane at its tip is implanted into a specific brain region. Small solutes including neurotransmitters, hormones, and drugs diffuse into the probe, where they can be collected for analysis. In this experiment, glucose concentration in the hippocampus was measured as a readout for local neural activity while rats completed a task requiring the use of spatial working memory.
You’ve just watched JoVE’s video on stereotaxic surgery. In this video, we’ve covered the principles and methods of the procedure as well as its many applications in neuroscience labs.
Thanks for watching!
View the full transcript and gain access to JoVE Science Education videos
Q1: What is a stereotaxic atlas and how is it used in brain surgery?
A stereotaxic atlas is a 3D reconstruction of the brain created from serial sections of stained brains. It provides spatial coordinates using three axes: anterior-posterior, medial-lateral, and dorsal-ventral. Researchers locate their target brain region in the atlas and use the grid to calculate corresponding coordinates, enabling precise targeting of deep brain structures during surgery.
Q2: How do surgeons identify anatomical landmarks like bregma and lambda?
Bregma and lambda are skull landmarks formed by intersections of bone plates called sutures. Bregma is where the sagittal suture meets the coronal suture, while lambda is more posterior where the sagittal suture intersects the lambdoid suture. These landmarks serve as reference points for measuring coordinates and positioning the surgical probe accurately during stereotaxic procedures.
Q3: What components make up a stereotaxic frame and what do they do?
The stereotaxic frame consists of a main frame, micromanipulator, and probe holder attached to a mounting clamp. Incisor and ear bars hold the animal's head in a fixed position. The micromanipulator enables precise movement of surgical probes to predetermined coordinates using sliding Vernier scales for exact measurements in all three dimensions.
Q4: How do you read a Vernier scale on a stereotaxic micromanipulator?
First, identify where the 0 line sits relative to tick marks on the right scale, which is labeled in 10 millimeter increments. Then find the line on the left scale that aligns best with a mark on the right side and note its value. Combine these readings to determine the precise coordinate measurement, such as 10.9 millimeters.
Q5: What are the main surgical steps for performing stereotaxic surgery?
After anesthetizing and positioning the animal, surgeons shave and disinfect the scalp, then expose the skull with a small incision. They level the brain by measuring at bregma and lambda, record coordinates, calculate probe movements to the target, drill a hole in the bone, and section the dura membrane. The micromanipulator then lowers the probe to the target depth for the desired manipulation.
Q6: How is stereotaxic surgery used to deliver genes or genetic material to the brain?
Genetically engineered viruses are injected into precise brain regions using stereotaxic surgery, delivering genes of interest to infected cells. Alternatively, viral delivery of small interfering RNA can suppress specific protein expression in targeted brain areas. This approach enables researchers to manipulate gene expression in discrete neural populations for studying cellular and molecular neuroscience research.
Q7: What recording and monitoring techniques use stereotaxic surgery to study brain function?
Stereotaxic surgery enables electrode placement for recording electrical signals from neurons during freely behaving tasks. Microdialysis probes with semipermeable membranes can be implanted to continuously monitor neurotransmitter, drug, or metabolite concentrations in awake animals. These techniques allow researchers to measure neural activity and brain chemistry while animals perform behavioral tasks requiring spatial working memory.
Chapters in this video
0:00
Overview
1:00
Principles of Stereotaxic Surgery and the Stereotaxic Frame
3:50
Stereotaxic Surgery
5:48
Applications
8:05
Summary
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