Study Suggests Disruptive Effects of Anesthesia on Brain Cell Connections Are Temporary

A study of juvenile rat brain cells suggests that the effects of a commonly used anesthetic drug on the connections between brain cells are temporary.

The study, published in this week’s issue of the journal PLOS ONE, was conducted by biologists at the University of California, San Diego and Weill Cornell Medical College in New York in response to concerns, arising from multiple studies on humans over the past decade, that exposing children to general anesthetics may increase their susceptibility to long-term cognitive and behavioral deficits, such as learning disabilities.

An estimated six million children, including 1.5 million infants, undergo surgery in the United States requiring general anesthesia each year and a least two large-scale clinical studies are now underway to determine the potential risks to children and adults.

“Since these procedures are unavoidable in most cases, it’s important to understand the mechanisms associated with the potentially toxic effects of anesthetics on the developing brain, and on the adult brain as well,” said Shelley Halpain, a professor of biology at UC San Diego and the Sanford Consortium for Regenerative Medicine, who co-headed the investigation. “Because the clinical studies haven’t been completed, preclinical studies, such as ours, are needed to define the effects of various anesthetics on brain structure and function.”

“There is concern now about cognitive dysfunction from surgery and anesthesia—how much these effects are either permanent or slowly reversible is very controversial,” said Hugh Hemmings, Jr., chair of anesthesiology at Weill Cornell and the study’s other senior author. “It has been suggested recently that some of the effects of anesthesia may be more lasting than previously thought. It is not clear whether the residual effects after an operation are due to the surgery itself, or the hospitalization and attendant trauma, medications and stress—or a combination of these issues.”

However, he added, “There is evidence that some of the delayed or persistent cognitive effects after surgery are not primarily due to anesthesia itself, but more importantly to brain inflammation resulting from the surgery. But this is not yet clear.”

The team of biologists examined one of the most commonly used general anesthetics, a derivative of ether called “isoflurane” used to maintain anesthesia during surgery.

“Previous studies in cultured neurons and in the intact brains of rodents provided evidence suggesting that exposure to anesthetics might render neurons more susceptible to cell death through a process called ‘apoptosis’,” said Halpain. “While overt cell death could certainly be one way to explain any long-lasting neurocognitive consequences of general anesthesia, we hypothesized that there could be other cellular mechanisms that disrupt neural circuits without inducing cell death per se.”

One such mechanism, she added, is known as “synaptotoxicity.” In this mechanism of neural-circuit disruption, the “synapses,” or junctions between neurons, become weakened or shrink away due to some factor that injures the neurons locally along their axons (the long processes of neurons that transmit signals) and dendrites (the threadlike extensions of neurons that receive nerve signals) without inducing the neurons themselves to die.

In the experiments at UC San Diego headed by Jimcy Platholi, a postdoctoral researcher in Halpain’s lab who is now at Weill Cornell, the scientists used neurons from embryonic rats taken from the hippocampus, a part of the mammalian forebrain essential for encoding newly acquired memories and ensuring that short-term memories are converted into long-term memories. The researchers cultured these brain cells in a laboratory dish for three weeks, allowing the neurons time to mature and to develop a dense network of synaptic connections and “dendritic spines”—specialized structures that protrude from the dendrites and are essential mediators of activity throughout neural networks.

“Evidence from animal studies indicates that new dendritic spines emerge and existing spines expand in size during learning and memory,” explained Halpain. “Therefore, the overall numbers and size of dendritic spines can profoundly impact the strength of neural networks. Since neural network activity underlies all brain function, changes in dendritic spine number and shape can influence cognition and behavior.”

Using neurons in culture, rather than intact animal brains, allowed the biologists to take images of the synapses at high spatial resolution using techniques called fluorescence light microscopy and confocal imaging. They also used time-lapse microscopy to observe structural changes in individual dendritic spines during exposure to isoflurane. Karl Herold, a research associate in the Hemmings laboratory and a co-author of the study, performed some of the image analysis.

“Imaging of human brain synapses at this level of detail is impossible with today’s technology and it remains very challenging even in laboratory rodents,” said Halpain. “It was important that we performed our study using rodent neurons in a culture dish, so that we could really drill down into the subcellular and molecular details of how anesthetics work.”

The researchers wondered whether brief exposure to isoflurane would alter the numbers and size of dendritic spines, so they applied the anesthetic to the cultured rat cells at concentrations and durations (up to 60 minutes) that are frequently used during surgery.

“We observed detectable decreases in dendritic spine numbers and shape within as little as 10 minutes,” said Halpain. “However this spine loss and shrinkage was reversible after the anesthetic was washed out of the culture.”

“Our study was reassuring in the sense that the effects are not irreversible and this fits in with known clinical effects,” said Hemmings. “For the most part, we find that the effects are reversible.”

“We clearly see an effect—a very marked effect on the dendritic spines—from use of this drug that was reversible, suggesting that it is not a toxic effect, but something more relevant to the pharmacological actions of the drug,” he added. “Connecting what we found to the cognitive effects of isoflurane will require much more detailed analysis.”

The team plans to follow up its study with future experiments to probe the molecular mechanisms and long-lasting consequences of isoflurane’s effects on neuron synapses and examine other commonly-used anesthetics for surgery.

How Anesthesia Works In Animals

There are basically two types of anesthetics: injectable anesthetics and gas anesthetics.  We at All Pets Veterinary Home Care use gas anesthesia nearly exclusively for our patients as they are typically both more consistent and safer than their injectable counterparts.  This means our patients and their owners can enjoy less risk & more comfort.

When an injectable anesthetic or sedative is injected into a patient, there is no way of getting it out once it’s in.  If the patient proves sensitive or allergic to the drug, we must simply do what we can to support them until the effects wear off.  Even in the rare case that there is an antidote to the drug, there is no guarantee that the patient will respond as desired or that we will be able to act fast enough for the patient to benefit from the antidote’s effects.  Of course, sometimes injectable anesthetics & sedatives are the best choice and in those cases we must simply be as cautious and prepared as possible in order to prevent and, if needed, handle any problems that may occur.

In those cases in which injectable drugs are not needed, we use isoflurane gas anesthesia.  This is a newer and much safer cousin to the ether gas used in the old days.  Isoflurane gas actually starts off as a liquid which is placed into a special machine called a vaporizer, turned into a gas and mixed with oxygen prior to being administered to a patient.  As the mixture of oxygen and isoflurane gas is breathed in by the patient, it is absorbed into the bloodstream through the lungs and travels to the brain where it does its work to place the patient into an anesthetic state.  The beauty of gas anesthesia is it works very quickly.  This not only means patients reach an anesthetic plane quickly but it also means that they can be woken up very quickly as well, either when the procedure being performed is complete OR if there is a problem of any kind during the procedure.  This is due to the fact that not only is the gas anesthesia delivered to the brain through the lungs but it also exits the body through the lungs.  This means that as soon as the anesthesia machine is turned OFF and no more anesthetic is being breathed IN, the patient begins breathing OUT all of the anesthesia that they have in their body.  With each and every breath, the patient has less and less anesthetic in their body and begins to wake up very quickly. 

This ability to control the anesthesia so precisely by using gas means there are less chances for complications to arise.  Contrast this with the injectable forms which often leave doctors and/or technicians sitting with patients for variable periods of time after a procedure while waiting for the anesthetic to wear off and your choice of anesthetic should be clear the next time your pet requires anesthesia! 

NOTE:  Of course, these statements are very general and, as everyone knows, we must be specific when dealing with the life of a patient.  No topic in medicine is black & white and it is the grey areas we must account for when formulating any diagnostic or treatment plan.  Therefore, every precaution must be taken and every complication must be ready to be dealt with should one occur at any time.  Only then can we be certain that we are doing what is best for our patients.

Does the future of antidepressants lie in anesthetics? 

Mood disorders like depression are among the leading causes of mental disability in the U.S. Because the therapeutic effects of traditional interventions and pharmacological treatments have shown to be limited in terms of efficacy, new therapies that are rapid acting and have increased efficacy in treatment-resistant populations are actively being sought out. Today, I was surprised to learn that the volatile anesthetic isoflurane has been shown to have antidepressant effects in humans since 1985 (Langer et al.) Given that I use isoflurane to knock my rats out while I implant cannulas and our lab studio the development of depressive-like behavior following early life abuse, I was floored

Although clinical literature has provided hints into the use of anesthetics, whether isoflurane could also exert antidepressant-like effects in conventional animal models of depression had not been determined until recently. Paul Shepard’s lab at the Maryland Psychiatric Research Center (MPRC) have behavioral evidence suggesting that a short period of acute isoflurane inhalation is sufficient to impede the development of depressive-likebehavior in rats. 

For this study, the group administered isoflurane (2% in 100% O2) to adult male Sprague Dawley rats continuously for two hours through a nose cone attached to a standard stereotaxic apparatus. Two weeks following exposure to isoflurane, rats entered a conventional two-day learned helplessness paradigm. Learned helplessness is a term that refers to the condition of an organism (human or animal) that has learned to behave helplessly and fails to respond even in the presence of opportunities for it to help itself.  Another way of thinking about it is as a perceived loss of control over the outcome of a situation on the organism’s part. Specifically, the group used the shuttle box avoidance task, in which an animal must move from one compartment to the other in order to gain either gain a reward or avoid an aversive stimulus such as a shock. 

As shown in the figure above, isoflurane-treated rats (n=12) had fewer failure trials (Fig. 1A) and a faster mean escape latency (Fig. 1B) in the shuttle box avoidance task compared to naïve-controls (n=12). To specify this effect, a separate group of rats was exposed to an equivalent dose (1.5% in 100% O2) of another anesthetic agent, halothane,  for two hours, and  evaluated in an identical learned helplessness paradigm after the same two week recovery period. Halothane-treated rats (n=12) performed similarly to naïve-controls (n=10; Fig. 1C-D), suggesting that the reduced expression of learned helplessness is specific to isoflurane rather than a general effect associated with exposure to volatile anesthetics. Importantly, these results support and extend on previous findings indicating that isoflurane has antidepressant effects in humans and provide new insights and opportunities regarding alternate targets for development of rapid pharmacological treatments for depression. 

Obviously, more work needs to be done in order to clarify the mechanism of action and the timing in which isoflurane inhalation could prove to be useful. For example, would isoflurane work if it were given during infancy or adolescence? Could it be sufficient to prevent the depressive-like behavior even in the face of early life adversity? Moreover, research designed at determining the minimum exposure of isoflurane necessary for this effect would also be useful. Would animals, like the clinical population exhibit individual differences in terms of what dose is effective?  We don’t know, but I certainly hope we find out. 

This work was presented on Tuesday, October 16th 2012 by L. Wang as a poster titled Isoflurane impedes the development of a depression-like phenotype in rats. 


For small animals:

Cats and dogs must be starved overnight before they come in however smaller
animals eg rabbits and guinea pigs should have food available until the
point of admission.
Pre-med is given in the morning before the operation which contains pain
relief and a sedative. This is given in the morning as it is more effective
if it is given before pain

Just before the operation, a intravenous injection containing Propofol
(milky white emulsion) is given into the foreleg (or ear for rabbits)
which is a short-term anaesthetic. NB Propofol is also used in people.

While the animal is unconscious, an endotracheal tube is put down their
trachea (easiest to put down when holding the animal upright and pulling
tongue down). There are a range of sizes going up in gaps of 0.5. A mixture
of oxygen and often isoflurane (for cats and dogs, I’m not sure about
rabbits) is pumped continuously through which continues to keep the animal

The machine that the endotracheal tube is hooked up to has a reservoir bag
(the size depends on the size of animal) which collects expired and fresh
gas during exhalation for the next breath. It also stops sudden pressure
changes and can be squeezed to cause inspiration manually.
It also has a CO2 absorber (I think it might contain soda lime but I’m not
sure) which absorbs any CO2 in the gases before the animal inhales them.
There is a control on the machine to change the percentage of isoflurane
(or equivalent) if the patient is too deeply anaesthetised or too awake.
Common signs of the animal not having enough anaesthetic are moving or
reacting to the operation (clearly), having a reflex when the eye is
touched (however I don’t think this is always reliable - particularly in
smaller animals and rodents), moving the paw when toe is pinched or
reacting when other sensitive skin is pinched. Another common method is
looking at the resistance when opening their jaw - if the animal appears to
be closing their jaw in response, they are probably too light.

Respiration and heart rate can be monitored throughout the operation using
an oesophageal stethoscope (I think) which goes down the oesophagus and
rests above the heart.

Whilst the animal is anaesthetised, there is a lot of heat loss
particularly for smaller animals (I assume because they have a larger
surface area:volume ratio). I’m not completely sure why - I have read that
it is because while unconscious animals cannot control their body
temperature and also that many anaesthetics affect the temperature control
in the brain and cause vasodilation. I don’t know how accurate this is but
it may be a combination of these. This means that heating mats and blankets
are used during the operation to ensure the dog remains warm.

That’s all I’ve learned for small animals so far but I’ll add anything new
I learn - this is just what I’ve picked up through watching operation and a
bit of research so I’m not completely sure everything is accurate!

Surgical Anesthetic Appears to Treat Drug-Resistant Depression

More study is needed, but isoflurane might provide alternative to electroconvulsive therapy

Although electroconvulsive therapy (ECT) has long been considered the most effective treatment of medication-resistant depression, millions of people who could benefit don’t take advantage of it because of the treatment’s side effects and public misperception of the procedure.

If the results of a campus-wide collaboration of University of Utah researchers are borne out by larger studies and trials, patients with refractory depression might one day have an alternative that is as effective as ECT but without the side effects – the surgical anesthetic drug isoflurane. 

“We need to expand our research into a larger, multicenter trial, but if the results of our pilot study pan out, it would change the face of treating depression,” says Howard R. Weeks, M.D., assistant professor of psychiatry and first author on a study published July 26, 2013, in PLOS ONE online.

Also known as shock therapy, ECT is effective in 55 percent to 90 percent of depression cases, with significant reductions in symptoms typically occurring within two to four weeks. When medications work, they can take six to eight weeks to become effective. But ECT is associated with side effects including amnesia, concentration and attention problems, and other cognitive issues. Many people also mistakenly believe ECT is painful and causes brain damage, which has given the treatment a social stigma that makes millions of patients reluctant to have it. Isoflurane potentially offers an alternative to ECT that could help many of those people, according to Weeks and his colleagues from eight University of Utah departments and programs. 

In a pilot study with 20 patients who received ECT treatments compared to eight patients who received the isoflurane treatments, the researchers found that both therapies provided significant reduction in symptoms of depression. Immediately following the treatments, ECT patients showed declines in areas of memory, verbal fluency, and processing speed. Most of these ECT-related deficits resolved by four weeks. However, autobiographical memory, or recall of personal life events, remained below pretreatment levels for ECT patients four weeks after the treatment. In contrast, the patients treated with isoflurane showed no real impairment but instead had greater improvements in cognitive testing than ECT patients both immediately and four weeks after the treatments. 

In the mid-1980’s, researchers in Europe studied isoflurane as a potential depression therapy. Later studies by other scientists failed to confirm the results of the original work and isoflurane research fell out of favor. But these later studies didn’t adhere to the first study’s protocol regarding type of anesthetic, dosing size and number of treatments, according to Weeks, and he believes that’s why isoflurane’s antidepressant effects weren’t confirmed in subsequent trials. For their research, Weeks and his University of Utah colleagues followed the original study’s protocol. 

“Our data reconfirm that isoflurane had an antidepressant effect approaching ECT with less adverse neurocognitive effects, and reinforce the need for a larger clinical trial,” the researchers wrote. 

Researchers don’t know what produces the relief of depression symptoms from ECT or isoflurane. Weeks believes further research might identify a molecular pathway that both therapies target and is responsible for the improvement in depression. One common effect of both ECT and isoflurane treatments is a brief state of low electrical activity in which the brain becomes unusually quiet. ECT induces a seizure to reach that state, but isoflurane does not. After inhaling the anesthesia, patients are “under” for about 45 minutes, with 15 minutes of that time being a deep state of unconsciousness, according to Weeks. This period of electrical rest for the brain may be a potential explanation for why ECT and isoflurane improve depression. 

If isoflurane proves to be a viable alternative to ECT, a device invented by three University of Utah anesthesiology faculty members can make the anesthetic an even more attractive therapy. The Anecleardevice (Anecare, Salt Lake City, UT) invented by Dwayne R. Westenskow, Ph.D., Derek J. Sakata, M.D., and Joseph A. Orr, Ph.D., from the University of Utah Department of Anesthesiology, uses hyperventilation and allows patients to rebreathe their own carbon dioxide (C02). Hyperventilation removes anesthesia from the lungs and C02 encourages blood flow to the brain, which encourages quicker removal of anesthetic. The Aneclearalso minimizes or even eliminates vomiting, nausea, and extreme fatigue that some patients experience from anesthesia. 

“With the Aneclear, we can wake people up from the anesthesia much quicker,” Weeks says. “This makes the treatment a potentially viable clinical treatment by reducing the time required in an operating room.” 

Weeks and his co-researchers now are looking for grants to fund a larger study that will include several U.S. centers.

Anesthetic Linked to Brain Cell Death in Newborn Mice

Exposure to the anesthetic agent isoflurane increases “programmed cell death” of specific types of cells in the newborn mouse brain, reports a study in the April issue of Anesthesia & Analgesia, official journal of the International Anesthesia Research Society (IARS).

With prolonged exposure, a common inhaled anesthesia eliminates approximately two percent of neurons in the cortex of newborn mice. Although its relevance to anesthesia in human newborns remains to be determined, the study by Dr George K. Istaphanous and colleagues of Cincinnati Children’s Hospital Medical Center provides unprecedented detail on the cellular-level effects of anesthetics on the developing brain.

Isoflurane Exposure Increases ‘Programmed Death’ of Brain Cells
In the study, seven-day-old mice were exposed to isoflurane for several hours. After exposure, sophisticated examinations were performed to assess the extent of isoflurane-induced brain cell death, including the specific types, locations, and functions of brain cells lost.

Isoflurane exposure led to widespread increases programmed cell death, called apoptosis, throughout the brain. Although cell loss was substantially higher after isoflurane exposure, the cell types lost were similar to the cells lost in the apoptosis that is part of normal brain maturation. In both cases, mainly neurons were lost. Neurons are the cells that transmit and store information.

The rate of cell death in the superficial cortex—the thick outer layer of the brain—was at least eleven times higher in isoflurane-exposed animals than seen with normal brain maturation. Overall, approximately two percent of cortical neurons were lost after isoflurane exposure. Astrocytes, another major type of cortical brain cells, were less affected by anesthetic exposure.

Relevance to Anesthesia in Human Newborns Is Unclear—For Now
A growing body of evidence suggests that isoflurane and similar anesthetics may have toxic effects on brain cells in newborn animals and humans. “However, neither the identity of dying cortical cells nor the extent of cortical cell loss has been sufficiently characterized,” according to Dr Istaphanous and colleagues.

The new study provides detailed information on the extent and types of brain cell loss resulting from prolonged isoflurane exposure in newborn mice. It’s unclear whether the two percent brain cell loss induced in the experiments would lead to any permanent damage—in previous studies, newborn isoflurane-exposed mice showed no obvious brain damage long after the exposure.

It can’t be assumed that isoflurane causes similar patterns of cellular damage in human newborns requiring general anesthesia, Dr Istaphanous and coauthors emphasize. Some studies have linked early-life exposure to anesthesia and surgery to later behavioral and learning abnormalities. Other studies have found no adverse affects on children exposed to anesthetics during vulnerable times of brain development. Further research on the selective nature and molecular mechanisms of isoflurane-induced brain cell death would be needed to determine the relevance of the experimental findings, if any, to human infants undergoing anesthesia.