18 September 2009

Cannabinoid Controversy

Activation of one of the subtypes of receptors binding cannabinoids (active ingredients of marijuana) suppresses inflammation, which makes this receptor a promising target for the treatment of autoimmune diseases such as multiple sclerosis and stroke complications that develop when tissues are damaged by immune cells. However, the fact that these receptors are present on neurons has caused heated debate among specialists, which can significantly reduce the potential of a new therapeutic target.

Traditionally, it is believed that the sensations caused by marijuana are based on the binding of cannabinoids to one of the types of CB1 cannabinoid receptors expressed in large quantities by brain neurons. Experts associate the functioning of these receptors with a number of problems, ranging from depression to obesity and cardiovascular diseases, as well as with therapeutic effects such as pain relief and nausea suppression. (The drug Marinol, a synthetic analogue of the active component of marijuana, was approved in 1985 to suppress nausea and improve appetite in patients with malignant diseases.) Despite the fact that a number of experts insist on the medicinal properties of cannabinoids, for the US Food and Drug Administration, the approval of compounds included in the No. 1 list (banned by the government on the basis of their danger to health and lack of justified medical use) is a risky step. (Although a hundred years ago, pharmacies freely sold, including such medicines as in this picture :)

About 10 years ago, scientists identified a second variant of cannabinoid receptors – CB2. They are found on immune cells and, apparently, play a role in the formation of the immune response. Until now, it was believed that CB2 receptors are practically absent on brain cells. This means that selective exposure to these receptors will avoid the psychotropic effects of cannabinoids and facilitate obtaining official approval. However, recently one of the research groups demonstrated evidence of the presence of a large number of CB2 receptors on the surface of neurons. Confirmation of this fact can greatly overshadow the hope of creating anti-inflammatory drugs that affect CB2 receptors and do not have undesirable side effects.

The group of Ron Tuma, a physiologist from Temple University (Philadelphia, Pennsylvania), is studying the role of CB2 receptors in the development of multiple sclerosis and stroke. CB2 receptors are expressed in large numbers by immune cells, and the scientists wanted to test whether their activation suppresses inflammation. In June of this year, they published an article in the journal Microvascular Research, according to which selective exposure to CB2 receptors significantly reduces the area of tissue damage and necrosis after stroke in mice.

At the same time, scientists from the University of Auckland, New Zealand, working under the guidance of pharmacologist Michelle Glass, have found that the activation of CB2 receptors is able to protect neurons from damage, most likely by preventing the activation of inflammatory processes by immune cells of the microenvironment of brain neurons or microglia.

The researchers hope that, due to the almost complete absence of CB2 receptors on neurons, drugs selectively binding to them will not have the psychotropic effects inherent in their analogues from list No. 1.

However, there is a fly in the ointment in this barrel. Microbiologist Emmanuel Onaivi from William Paterson University (Wayne, New Jersey) and his colleagues last year demonstrated that the administration of CB2 receptor agonists to stressed mice increases their cravings for alcohol. In 2006, the same author published data according to which Japanese men with a certain variant of the CB2 receptor gene are characterized by a high probability of developing alcoholism and depression. The results of both studies indicate the probable presence of CB2 receptors on brain cells. However, the most controversial data were published by Onaivi in 2006 in Brain and Research and in 2008 in the New York Academy of Sciences. Using labeled antibodies, Onaivi's group revealed the ubiquitous presence of CB2 receptors on neurons. In general, the results obtained by Onaivi indicate that selective agonists of CB2 receptors can act on the psyche in the same way as cannabinoids themselves, which makes their clinical use impossible.

It is a little encouraging that none of the laboratories were able to reproduce the results of Onaivi. Many researchers have unsuccessfully used specific antibodies to detect informational RNAs or active genes in neurons that provide expression of CB2 receptors.

Onaivi states that his predecessors were unable to detect the expression of CB2 receptors on neurons due to the imperfection of the methods of Western blotting and studying gene expression used by them. Other researchers disagree with this and believe that Onaivi himself could not use any fundamentally different approaches and that, on the contrary, his work cannot be considered adequately performed due to the lack of a well-thought-out set of controls.

For example, Onaivi identified various variants of CB2 receptors on the brain cells of wild-type mice, but used only one control group of "knocked out" animals without CB2 receptors. In addition, to identify CB2 receptors, he used peroxidase, which quite often gives false positive results. The longer sections of brain tissue are incubated in peroxidase, the more likely it is that a stain indicating the presence of receptors on the cells will appear. Such false-positive results can appear even when staining mouse tissue samples that do not have CB2 receptors.

According to Ken Mackie, who studies cannabinoid receptors at Indiana State University (Bloomington), when conducting such experiments, it is important not only parallel processing and analysis of experimental and control samples, but also the use of the "blind method" when the performer does not know which sample is in his hands. Onaivi did not adhere to these principles, so the reliability of the results obtained by his group is highly questionable. Moreover, in addition to detecting CB2 receptors on the cell surface, it is necessary to demonstrate the presence inside the cells of a large number of their constituent proteins and their corresponding informational RNAs.

However, Onaivi has an alternative explanation for the fact that he was able to detect a large number of receptors where others saw only traces of them. He claims that with the help of genomic analysis he proved the existence of two different isoforms of CB2 receptors. Preliminary results of this work were published in June in the journal Genes, Brain and Behavior. One of the isoforms is predominantly expressed in the brain and seminal glands, while the other is in the spleen, where CB2 receptors were first detected. Onaivi believes that the antibodies used by other researchers do not react with the CB2 receptor isoform expressed in the brain.

The existence of several isoforms of the same receptor encoded by different genetic sequences is quite likely, and to date no one can either confirm or refute Onaivi's statement. Meanwhile, the fate of a new class of anti-inflammatory drugs hung in the air indefinitely.

Evgeniya Ryabtseva
Portal "Eternal youth" http://vechnayamolodost.ru based on the materials of The Scientist: Cannabinoid controversy.


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