HÉLIO DE ALMEIDASuddenly, the heart starts thumping. One feels short of air, and is overcome by the terrifying certainty that one is going to die. Half an hour later, just as unexpectedly as when it came on, without any apparent reason, this state of extreme anxiety disappears. After the first crisis, which usually arises between the end of adolescence and the age of 40, certainty and security fade away, as if, from one moment to the next, a world of tranquility had crumbled. The person therefore lives under the constant threat of another sudden attack, which could happen at any time or in any place.
The chemical and biological transformations that spark off and, at the same time, feed these two alterations in the emotions – the blander, anxiety, and the deeper, a crisis of panic – are now far better understood and can be combated more efficiently, as a result of the studies on two substances, glutamate and nitric oxide, carried out by researchers from the School of Medicine of the University of São Paulo in Ribeirão Preto.
In experiments with rats and on human beings, the team led by a doctor from Rio Grande do Sul, Francisco Silveira Guimarães, proved the involvement of glutamate and nitric oxide, whose functions in the nervous system were as yet little known, in the manifestations of anxiety, panic, and in another emotional disorder as well, depression, characterized by a feeling of deep and persistent despondency and sadness. Before, there had only been indications of this participation, but not a definition of the specific role that they play in each problem.
Guimarães confirmed for the first time the participation of nitric oxide in anxiety, a disorder that affects 4% of the Brazilian adult population – almost 5 million persons in the country have shown at least one clinically identified episode of anxiety, marked in one of its most flagrant signs by an exaggerated concern with mundane facts (is the money going to run out before the end of the month? I am going to manage to do all the things that I have got to do?). The team from Ribeirão Preto found that the two substances, acting in conjunction with others, set in motion the mechanisms that lead to the panic disorder, a disturbance in which individuals show more than one crisis a week.
It is estimated that about 1.6% of the Brazilians have shown panic disorder at least once during their lifetime, according to a study coordinated by Laura Andrade, from USP’s Psychiatry Institute, and published in July 2002 in Social Psychiatry and Psychiatric Epidemiology. The work shows also shows that, for reasons that are not yet totally understood, women are 2.3 times more inclined than men to develop anxiety and panic, and 1.6 times more inclined to have depression.
Communication between neurons
In the organism, glutamate and nitric oxide act as chemical messengers – they are the so-called neurotransmitters -, carrying information from one nerve cell (neuron) to another. It is on this communication between neurons that the whole workings of the organism depend, from thought to conscious actions, like the movements of the hand of someone playing an instrument, to involuntary processes like breathing. In the central nervous system, made up of the brain and associated organs, like the encephalic trunk, the cerebellum and the spinal cord – jointly responsible for the general maintenance of the organism -, there are other neurotransmitters, serotonin, noradrenaline and gamma aminobutyric acid (Gaba), better studied, which also influence the workings of the neurons. The lack or excess of any one of them alters emotional well-being.
The results of the studies at USP in Ribeirão Preto facilitate, in first place, an understanding of the mechanisms by which some medicines act to increase the quantity of serotonin in the central nervous system, one of the strategies most adopted nowadays to combat anxiety, panic and depression, the most common psychic problems these days. Among the ones most used is the drug called fluoxetine, the basis for the famous Prozac, launched in 1986, which makes this neurotransmitter stay in action longer before degrading itself.
By showing how and where glutamate and nitric oxide act, the work by Guimarães’s team opens up new prospects for developing other drugs that, in future, may be used in association with the current ones in therapy for anxiety and depression, which afflict 16% of the Brazilians, according to Laura’s study. Medicines that act on glutamate or nitric acid could also be an alternative to the existing antidepressives, which mitigate the problem in a few weeks. What is undeniable is that they bring about a better quality of life, particularly when associated with long term psychotherapeutic monitoring, focusing on a search for the deeper causes of these disorders and for new ways of dealing with day to day problems.
Medicines alone are not a definitive cure for these emotional upsets, which, in the view of the psychoanalysts, are problems that are typical of the last few decades, a period in which there is a prevalence of values such as individualism, consumerism, and a giddy succession of events that pile up, as if each day were too short for the planned tasks. “There is a tendency for these emotional alterations to arise in more individualist societies, where people have fewer guarantees assured them by culture and by social ties,” explains psychiatrist and psychoanalyst Mário Eduardo Pereira, the director of the Fundamental Psychopathology Laboratory of the Campinas State University (Unicamp). “As a consequence, the forlornness is experienced more intensely.”
Anxiety, panic and depression could thus be a result of a kind of being out of step between the changes in life (less time to see friends and more work, for example) and the capacity of human beings to adapt to them, a view that is shared by Márcio Giovannetti, the president of the Brazilian Psychoanalysis Society of São Paulo (SBPSP). Ana Maria Sigal, a psychoanalyst and professor of psychoanalysis at the Sedes Sapientiae Institute, in São Paulo, observes daily the effects of the contemporary way of life. “We psychoanalysts are finding today a considerable increase of patients who complain of crises of anxiety and panic” says she.
Although the specific causes of anxiety, panic and depression are still not very well defined, doctors and psychoanalysts are in agreement: these disorders arise as a consequence of a combination of three factors. Firstly, there are the biological fuses, that is, the genetic predisposition for suffering from one of these disorders at some moment in life. The emotional factors follow – people who are more vulnerable or more sensitive to the facts of reality tend to be more habitual victims of anxiety and depression. Finally, the environmental reasons, like difficulty in adapting to the transformations of society. “There isn’t a single cause for these emotional problems, but an integration of these three factors,” psychologist Mariângela Gentil Savoia, from the Anxiety Clinic of USP’s Psychiatry Institute explains. “Just one of the factors, in isolations, is not sufficient for the disorder to manifest itself.”
In a study of 43 victims of panic disorder and another 29 healthy volunteers, Mariângela assessed the number of stressful situations that the participants had experienced in the year prior to the first crisis, and found that what varied was not the number of events, but the way that people dealt with them and the value they attributed to them. The main one of these factors was the loss of social support, or, more precisely, of a relative or a friend, one year before the first panic crisis. “People who panic show strategies that are not very well adapted to face up to adverse situations'” says the psychologist. “Generally speaking, they do not try to solve the problem, they avoid it rather.” That is why the use of medicines may not be the definitive solution, but it does help the patient to face up to the treatment that combines medicine and psychotherapy.
It so happens that to get a medicine that acts in a selective way on glutamate or nitric oxide and causes fewer collateral effects still needs years of research, Guimarães warns. But some of these new drugs are now appearing. One of them is memantine, which seems to act on a molecule to which glutamate binds itself, and so inhibits its activity. Produced since 1989 by the German laboratory Merz, it was presented again in Europe in October of last year, with a new purpose: to fight Alzheimer, a disease that causes the degeneration of the central nervous system and the loss of memory. Perhaps, in future, memantine or similar medicines may help to resolve anxiety and panic too, and even depression, with the advantage of not causing hallucinations, like another drug used only experimentally, AP-7, an abbreviation for 2-amino-7-phosphono-heptanoic acid.
Overprotective families
Even this ideal drug, which acts on glutamate or nitric acid, is not sufficient. “In an isolated way, no medicine resolves panic, anxiety and the phobias,” Giovannetti, from the SBPSP, observes. “Medicines help, but they do not modify the essence that generates the problem, because man is a biological, psychic and social being. The existence of each one of us does not react only to organic factors,” the psychoanalyst comments. Psychiatrist Mário Eduardo Pereira arrived at a conclusion similar to Giovanetti’s during the treatment of victims of panic disorder at Unicamp. “Generally speaking, the medicines were useful for controlling the crises, but that was insufficient for the clinical treatment of these individuals'” he explains. “Many patients were afraid of starting to use the drug; others, when they stopped the treatment, suffered from new crises, which created the need for continuous use of the medicine.”
Eager to understand the problem better, in 1995 Pereira embarked on a doctorate in psychoanalysis at the Paris VII University, in France. When assessing victims of panic disorder attended to at the university between 1984 and 1995, now from the point of view of psychoanalysis, he found that two distinct groups predominated: those who came from overprotective families, who had always lived in a secure environment, without ever having actually faced up on their own to the reality of the lack of guarantees for existence; and another, with opposite characteristics, members of families who looked on the day to day facts as terrifying.
“It was then we started to understand that, from the clinical point of view, treatment with medicine only makes sense when you have a broader view of the individual,” Pereira comments. “You have to know how the crises arise and what are the symbolic and personal dimensions involved in their lives, in connection with the attacks.”
Cats and depression
Conclusions like these are united in the search for solutions that include new medicines and an understanding of the biological mechanisms for anxiety, panic and depression. In Francisco Guimarães’s laboratory at USP in Ribeirão Preto, it is common to see technicians injecting medicines directly into the brains of rats, or rats walking in suspended labyrinths or cages, face to face with cats. These were some of the resources of work that made it possible for the research group to assess the alterations in the rodents’ behavior after glutamate and nitric oxide were injected in areas of the central nervous system associated with fear, such as the dorsal periaqueductal gray matter, a set of neurons located in the encephalic trunk, between the brain and the spinal cord.
The first clues that led Guimarães to study glutamate and, afterwards, nitric oxide, arose in the second half of the 80’s. At the time, the Brazilian researcher was studying for a doctorate as a student of psychiatrist Antônio Zuardi and psychopharmacologist Frederico Graeff, who had demonstrated that serotonin, when applied directly into the periaqueductal gray, would diminish the responses of anxiety, when facing situations that awakened fear. “It remained to find out which neurotransmitter was acting in the stimulation of this region”, Guimarães explains.
One of the candidates was glutamate, the main stimulant chemical messenger in the central nervous system. Richard Bandler, in Australia, and Graeff himself observed that a mere injection of glutamate, without any exposure to a threatening situation, would produce manifestations that seemed to be associated with reactions of panic in humans. But these indications were not enough to prove that the alteration observed in the behavior of the animals derived from the action of glutamate. It was still not possible to know whether the level of glutamate in this region of the central nervous system would increase, in situations that create anxiety and panic,” the researcher explains. In a study in collaboration with Graeff, José Carlos de Aguiar and Antonio de Padua Carobrez, today a researcher at the Federal University of Santa Catarina (UFSC), Guimarães used AP-7, which inhibits the action of glutamate, and has been employed for almost 20 years in tests with laboratory animals.
They put the little rats into a piece of equipment called a cross-shaped maze, an X-shaped platform, 50 centimeters from the ground, with two arms open and another two protected by walls. As they usually are afraid of height and open spaces, the rats seek the closed parts, rather like what happens with someone who is afraid of heights and is put on the balcony of a building. They found that the rodents treated with AP-7 seemed to have lost their fear: they would leave the protected areas and visit the open areas of the maze two times more, compared with rats that had been given an injection with only water and salt – the result was a confirmation of the participation of glutamate as a stimulant for anxiety: “Had this neurotransmitter not exercised an active physiological role in anxiety, the rats would have behaved similarly, avoiding going out into the open regions” Guimarães claims. “The treated rats lost their fear, and some of them even went so far as to fall off the platform'” he comments.
The proofs
In 1991, English pharmacologist John Garthwaite, from University College, in London, suggested, in a study published in Trends in Neuroscience, that the action of glutamate in the brain could be, in part, a result of the production of nitric oxide – a gas that not only acts as a chemical messenger in central nervous system, but also as a dilator of the blood vessels in other regions of the body. In the following year, the team led by Steven Vincent, from the University of British Colombia, in Canada, took a step forward and, in an indirect manner – through the detection of an enzyme that produces this neurotransmitter, nitric oxide synthase – mapped the regions of the central nervous system in which this gas acted. There, one more clue was to be found: in the dorsal periaqueductal gray matter, the enzyme was present in a large quantity – an indication of the participation of nitric oxide in manifestations of anxiety and panic.
With this information in hand, Guimarães and another two researchers from USP in Ribeirão, Elaine Del Bel and Gustavo Ballejo, applied in the periaqueductal gray matter of rats other compounds that inhibited the action of the nitric oxide producing enzyme. To prove the influence of this neurotransmitter gas, though, it had to be seen whether the opposite also happens, that is, when the quantity of nitric oxide is increased in the periaqueductal gray matter, reactions of anxiety and panic are induced.
Rúbia Weffort de Oliveira, who is studying for a doctorate under Guimarães, injected into the periaqueductal gray matter of rodents different doses of two compounds that release nitric oxide – 3-morpholinosydnonimine hydrochloride and a diethylamine/nitric oxide complex. Next, she put each one of the animals in isolation into an arena with 40-centimeter high plastic walls, and she found that the larger the dosage of the compounds, the more intense was the reaction of the rats. With higher doses, the rats would rush off running in circles and, in a desperate measure, would try – and sometimes succeed – to escape from the arena. Rúbia also found that if she treated the rats with methylene blue, a compound with an opposite effect to the two medicines, the animals would not show any signs of anxiety nor of panic.
In this work, published in 2000 in the Brain Research Bulletin, the researchers also succeeded in mapping the areas that were stimulated by the nitric oxide. By detecting a protein present in a greater quantity only in active nerve cells, they saw that the increase of nitric oxide in the periaqueductal region would stimulate neurons, not only in this area, but also in other parts of the central nervous system connected with the circuit of fear and anxiety, like the tonsils, responsible for remembering disagreeable events, and the hypothalamus, a center that controls neurovegetative reactions like breathing, the heartbeats, and the flight behavior.
What was missing, though, was to confirm if situations of stress really did increase the production of the nitric oxide producing enzyme. When immobilizing the rodents in cages, in a situation of extreme stress for rats, they verified an increase in the number of neurons that produced this enzyme in the periaqueductal region, according to an article published last year in Neuroscience and Biobehavioral Reviews. This result supplemented the work that had recently been published in the NeuroReport magazine by Silvana Chiavegatto, Cristoforo Scavone and Newton Canteras, from USP’s Medical Sciences Institute, showing that the exposure of rats to a cat increased the production of nitric oxide in this region of the central nervous system.
Shocks and flight
In parallel with these experiments, Guimarães investigated whether glutamate, the main central nervous system stimulating neurotransmitter, could influence the appearance of depression, an alteration in the emotions with characteristics practically opposite to those of anxiety. The suspicion was that, in a similar way to what happens in the periaqueductal region – in which serotonin and glutamate have opposite functions (the former inhibits the circuits of fear and anxiety, while the latter stimulates them) -, the same was to be verified in the hippocampus, an area of the brain connected with the memory of disagreeable events, which cause stress and the consequent paralysis of the animal.
In another experiment, the rats would receive electric shocks in their paws and, on the following day, they would first hear an audible signal and then feel the shock, so that the possibility of flight existed. As stress normally increases the level of glutamate in the hippocampus, the animals would not budge. Cláudia Padovan, who is studying for her doctorate, found that the rats would learn to flee after the application in the hippocampus of a substance that inhibits the action of glutamate.
Sâmia Joca, another student for a doctorate under Guimarães, found effects similar to those with the glutamate inhibitor when she injected the animals with zimelidine, a medicine similar to fluoxetine, which increases the quantity of serotonin in the central nervous system. “In the hippocampus, serotonin seems to assuage the emotional impact of stressful events, facilitating adaptation to stress and combating depression, while glutamate could have the opposite effect”, says Guimarães. His plan is to set off now for more refined studies and to assess the action of nitric oxide and glutamate in other regions of the nervous system associated with panic and anxiety.
The project
Neurobiology of Behavioral Responses to Aversive Events (98/10639-7); Modality: Thematic project; Coordinator: Francisco Silveira Guimarães – College of Medicine at Ribeirão Preto/USP; Investment: R$ 278,831.34 and US$ 129,952.32
