What is Midazolam?

Introduction

Midazolam, sold under the brand name Versed, among others, is a benzodiazepine medication used for anaesthesia, procedural sedation, trouble sleeping, and severe agitation.

It works by inducing sleepiness, decreasing anxiety, and causing a loss of ability to create new memories. It is important to note that this drug does not cause an individual to become unconscious, merely be sedated. It is also useful for the treatment of seizures. Midazolam can be given by mouth, intravenously, by injection into a muscle, by spraying into the nose, or through the cheek. When given intravenously, it typically begins working within five minutes; when injected into a muscle, it can take fifteen minutes to begin working. Effects last between one and six hours.

Side effects can include a decrease in efforts to breathe, low blood pressure, and sleepiness. Tolerance to its effects and withdrawal syndrome may occur following long-term use. Paradoxical effects, such as increased activity, can occur especially in children and older people. There is evidence of risk when used during pregnancy but no evidence of harm with a single dose during breastfeeding. It belongs to the benzodiazepine class of drugs and works by increasing the activity of the GABA neurotransmitter in the brain.

Midazolam was patented in 1974 and came into medical use in 1982. It is on the World Health Organisation’s List of Essential Medicines. Midazolam is available as a generic medication. In many countries, it is a controlled substance.

Brief History

Midazolam is among about 35 benzodiazepines currently used medically, and was synthesized in 1975 by Walser and Fryer at Hoffmann-LaRoche, Inc in the United States. Owing to its water solubility, it was found to be less likely to cause thrombophlebitis than similar drugs. The anticonvulsant properties of midazolam were studied in the late 1970s, but not until the 1990s did it emerge as an effective treatment for convulsive status epilepticus. As of 2010, it is the most commonly used benzodiazepine in anaesthetic medicine. In acute medicine, midazolam has become more popular than other benzodiazepines, such as lorazepam and diazepam, because it is shorter lasting, is more potent, and causes less pain at the injection site. Midazolam is also becoming increasingly popular in veterinary medicine due to its water solubility. In 2018 it was revealed the CIA considered using Midazolam as a “truth serum” on terrorist suspects in project “Medication”.

Medical Uses

Seizures

Midazolam is sometimes used for the acute management of seizures. Long-term use for the management of epilepsy is not recommended due to the significant risk of tolerance (which renders midazolam and other benzodiazepines ineffective) and the significant side effect of sedation. A benefit of midazolam is that in children it can be given in the cheek or in the nose for acute seizures, including status epilepticus. Midazolam is effective for status epilepticus that has not improved following other treatments or when intravenous access cannot be obtained, and has advantages of being water-soluble, having a rapid onset of action and not causing metabolic acidosis from the propylene glycol vehicle (which is not required due to its solubility in water), which occurs with other benzodiazepines.

Drawbacks include a high degree of breakthrough seizures – due to the short half-life of midazolam – in over 50% of people treated, as well as treatment failure in 14-18% of people with refractory status epilepticus. Tolerance develops rapidly to the anticonvulsant effect, and the dose may need to be increased by several times to maintain anticonvulsant therapeutic effects. With prolonged use, tolerance and tachyphylaxis can occur and the elimination half-life may increase, up to days. There is evidence buccal and intranasal midazolam is easier to administer and more effective than rectally administered diazepam in the emergency control of seizures.

Procedural Sedation

Intravenous midazolam is indicated for procedural sedation (often in combination with an opioid, such as fentanyl), for preoperative sedation, for the induction of general anaesthesia, and for sedation of people who are ventilated in critical care units. Midazolam is superior to diazepam in impairing memory of endoscopy procedures, but propofol has a quicker recovery time and a better memory-impairing effect. It is the most popular benzodiazepine in the intensive care unit (ICU) because of its short elimination half-life, combined with its water solubility and its suitability for continuous infusion. However, for long-term sedation, lorazepam is preferred due to its long duration of action, and propofol has advantages over midazolam when used in the ICU for sedation, such as shorter weaning time and earlier tracheal extubation.

Midazolam is sometimes used in neonatal intensive care units. When used, additional caution is required in newborns; midazolam should not be used for longer than 72 hours due to risks of tachyphylaxis, and the possibility of development of a benzodiazepine withdrawal syndrome, as well as neurological complications. Bolus injections should be avoided due to the increased risk of cardiovascular depression, as well as neurological complications. Midazolam is also sometimes used in newborns who are receiving mechanical ventilation, although morphine is preferred, owing to its better safety profile for this indication.

Sedation using midazolam can be used to relieve anxiety and manage behaviour in children undergoing dental treatment.

Agitation

Midazolam, in combination with an antipsychotic drug, is indicated for the acute management of schizophrenia when it is associated with aggressive or out-of-control behaviour.

End of Life Care

In the final stages of end-of-life care, midazolam is routinely used at low doses via subcutaneous injection to help with agitation, myoclonus, restlessness or anxiety in the last hours or days of life. At higher doses during the last weeks of life, midazolam is considered a first line agent in palliative continuous deep sedation therapy when it is necessary to alleviate intolerable suffering not responsive to other treatments, but the need for this is rare.

Administration

Routes of administration of midazolam can be oral, intranasal, buccal, intravenous, and intramuscular.

  • Dosing:
    • Perioperative use: 0.15 to 0.40 mg/kg IV.
    • Premedication: 0.07 to 0.10 mg/kg IM.
    • Intravenous sedation: 0.05 to 0.15 mg/kg IV.

Contraindications

Benzodiazepines require special precaution if used in the elderly, during pregnancy, in children, in alcohol- or other drug-dependent individuals or those with comorbid psychiatric disorders. Additional caution is required in critically ill patients, as accumulation of midazolam and its active metabolites may occur. Kidney or liver impairments may slow down the elimination of midazolam leading to prolonged and enhanced effects. Contraindications include hypersensitivity, acute narrow-angle glaucoma, shock, hypotension, or head injury. Most are relative contraindications.

Side Effects

Refer to Long-Term Effects of Benzodiazepines.

Side effects of midazolam in the elderly are listed above. People experiencing amnesia as a side effect of midazolam are generally unaware their memory is impaired, unless they had previously known it as a side effect.

Long-term use of benzodiazepines has been associated with long-lasting deficits of memory, and show only partial recovery six months after stopping benzodiazepines. It is unclear whether full recovery occurs after longer periods of abstinence. Benzodiazepines can cause or worsen depression. Paradoxical excitement occasionally occurs with benzodiazepines, including a worsening of seizures. Children and elderly individuals or those with a history of excessive alcohol use and individuals with a history of aggressive behaviour or anger are at increased risk of paradoxical effects. Paradoxical reactions are particularly associated with intravenous administration. After night-time administration of midazolam, residual ‘hangover’ effects, such as sleepiness and impaired psychomotor and cognitive functions, may persist into the next day. This may impair the ability of users to drive safely and may increase the risk of falls and hip fractures. Sedation, respiratory depression and hypotension due to a reduction in systematic vascular resistance, and an increase in heart rate can occur. If intravenous midazolam is given too quickly, hypotension may occur. A “midazolam infusion syndrome” may result from high doses, and is characterised by delayed arousal hours to days after discontinuation of midazolam, and may lead to an increase in the length of ventilatory support needed.

In susceptible individuals, midazolam has been known to cause a paradoxical reaction, a well-documented complication with benzodiazepines. When this occurs, the individual may experience anxiety, involuntary movements, aggressive or violent behaviour, uncontrollable crying or verbalization, and other similar effects. This seems to be related to the altered state of consciousness or disinhibition produced by the drug. Paradoxical behaviour is often not recalled by the patient due to the amnesia-producing properties of the drug. In extreme situations, flumazenil can be administered to inhibit or reverse the effects of midazolam. Antipsychotic medications, such as haloperidol, have also been used for this purpose.

Midazolam is known to cause respiratory depression. In healthy humans, 0.15 mg/kg of midazolam may cause respiratory depression, which is postulated to be a central nervous system (CNS) effect. When midazolam is administered in combination with fentanyl, the incidence of hypoxemia or apnoea becomes more likely.

Although the incidence of respiratory depression/arrest is low (0.1-0.5%) when midazolam is administered alone at normal doses, the concomitant use with CNS acting drugs, mainly analgesic opiates, may increase the possibility of hypotension, respiratory depression, respiratory arrest, and death, even at therapeutic doses. Potential drug interactions involving at least one CNS depressant were observed for 84% of midazolam users who were subsequently required to receive the benzodiazepine antagonist flumazenil. Therefore, efforts directed toward monitoring drug interactions and preventing injuries from midazolam administration are expected to have a substantial impact on the safe use of this drug.

Pregnancy and Breastfeeding

Midazolam, when taken during the third trimester of pregnancy, may cause risk to the neonate, including benzodiazepine withdrawal syndrome, with possible symptoms including hypotonia, apnoeic spells, cyanosis, and impaired metabolic responses to cold stress. Symptoms of hypotonia and the neonatal benzodiazepine withdrawal syndrome have been reported to persist from hours to months after birth. Other neonatal withdrawal symptoms include hyperexcitability, tremor, and gastrointestinal upset (diarrhoea or vomiting). Breastfeeding by mothers using midazolam is not recommended.

Elderly

Additional caution is required in the elderly, as they are more sensitive to the pharmacological effects of benzodiazepines, metabolise them more slowly, and are more prone to adverse effects, including drowsiness, amnesia (especially anterograde amnesia), ataxia, hangover effects, confusion, and falls.

Tolerance, Dependence, and Withdrawal

A benzodiazepine dependence occurs in about one-third of individuals who are treated with benzodiazepines for longer than 4 weeks, which typically results in tolerance and benzodiazepine withdrawal syndrome when the dose is reduced too rapidly. Midazolam infusions may induce tolerance and a withdrawal syndrome in a matter of days. The risk factors for dependence include dependent personality, use of a benzodiazepine that is short-acting, high potency and long-term use of benzodiazepines. Withdrawal symptoms from midazolam can range from insomnia and anxiety to seizures and psychosis. Withdrawal symptoms can sometimes resemble a person’s underlying condition. Gradual reduction of midazolam after regular use can minimise withdrawal and rebound effects. Tolerance and the resultant withdrawal syndrome may be due to receptor down-regulation and GABAA receptor alterations in gene expression, which causes long-term changes in the function of the GABAergic neuronal system.

Chronic users of benzodiazepine medication who are given midazolam experience reduced therapeutic effects of midazolam, due to tolerance to benzodiazepines. Prolonged infusions with midazolam results in the development of tolerance; if midazolam is given for a few days or more a withdrawal syndrome can occur. Therefore, preventing a withdrawal syndrome requires that a prolonged infusion be gradually withdrawn, and sometimes, continued tapering of dose with an oral long-acting benzodiazepine such as clorazepate dipotassium. When signs of tolerance to midazolam occur during intensive care unit sedation the addition of an opioid or propofol is recommended. Withdrawal symptoms can include irritability, abnormal reflexes, tremors, clonus, hypertonicity, delirium and seizures, nausea, vomiting, diarrhoea, tachycardia, hypertension, and tachypnoea. In those with significant dependence, sudden discontinuation may result in withdrawal symptoms such as status epilepticus that may be fatal.

Overdose

Refer to Benzodiazepine Overdose.

A midazolam overdose is considered a medical emergency and generally requires the immediate attention of medical personnel. Benzodiazepine overdose in healthy individuals is rarely life-threatening with proper medical support; however, the toxicity of benzodiazepines increases when they are combined with other CNS depressants such as alcohol, opioids, or tricyclic antidepressants. The toxicity of benzodiazepine overdose and risk of death is also increased in the elderly and those with obstructive pulmonary disease or when used intravenously. Treatment is supportive; activated charcoal can be used within an hour of the overdose. The antidote for an overdose of midazolam (or any other benzodiazepine) is flumazenil. While effective in reversing the effects of benzodiazepines it is not used in most cases as it may trigger seizures in mixed overdoses and benzodiazepine dependent individuals.

Symptoms of midazolam overdose can include:

  • Ataxia.
  • Dysarthria.
  • Nystagmus.
  • Slurred speech.
  • Somnolence (difficulty staying awake).
  • Mental confusion.
  • Hypotension.
  • Respiratory arrest.
  • Vasomotor collapse.
  • Impaired motor functions:
    • Impaired reflexes.
    • Impaired coordination.
    • Impaired balance.
    • Dizziness.
  • Coma.
  • Death.

Detection in Body Fluids

Concentrations of midazolam or its major metabolite, 1-hydroxymidazolam glucuronide, may be measured in plasma, serum, or whole blood to monitor for safety in those receiving the drug therapeutically, to confirm a diagnosis of poisoning in hospitalised patients, or to assist in a forensic investigation of a case of fatal overdosage. Patients with renal dysfunction may exhibit prolongation of elimination half-life for both the parent drug and its active metabolite, with accumulation of these two substances in the bloodstream and the appearance of adverse depressant effects.

Interactions

Protease inhibitors, nefazodone, sertraline, grapefruit, fluoxetine, erythromycin, diltiazem, clarithromycin inhibit the metabolism of midazolam, leading to a prolonged action. St John’s wort, rifapentine, rifampin, rifabutin, phenytoin enhance the metabolism of midazolam leading to a reduced action. Sedating antidepressants, antiepileptic drugs such as phenobarbital, phenytoin and carbamazepine, sedative antihistamines, opioids, antipsychotics and alcohol enhance the sedative effects of midazolam. Midazolam is metabolised almost completely by cytochrome P450-3A4. Atorvastatin administration along with midazolam results in a reduced elimination rate of midazolam. St John’s wort decreases the blood levels of midazolam. Grapefruit juice reduces intestinal 3A4 and results in less metabolism and higher plasma concentrations.

Pharmacology

Midazolam is a short-acting benzodiazepine in adults with an elimination half-life of 1.5-2.5 hours. In the elderly, as well as young children and adolescents, the elimination half-life is longer. Midazolam is metabolised into an active metabolite alpha1-hydroxymidazolam. Age-related deficits, renal and liver status affect the pharmacokinetic factors of midazolam as well as its active metabolite. However, the active metabolite of midazolam is minor and contributes to only 10 percent of biological activity of midazolam. Midazolam is poorly absorbed orally, with only 50% of the drug reaching the bloodstream. Midazolam is metabolised by cytochrome P450 (CYP) enzymes and by glucuronide conjugation. The therapeutic as well as adverse effects of midazolam are due to its effects on the GABAA receptors; midazolam does not activate GABAA receptors directly but, as with other benzodiazepines, it enhances the effect of the neurotransmitter GABA on the GABAA receptors (↑ frequency of Cl- channel opening) resulting in neural inhibition. Almost all of the properties can be explained by the actions of benzodiazepines on GABAA receptors. This results in the following pharmacological properties being produced: sedation, induction of sleep, reduction in anxiety, anterograde amnesia, muscle relaxation and anticonvulsant effects.

Pharmacokinetics

  • Volume of Distribution: 1-2.5L/kg in normal healthy individuals.
  • Protein Binding: 96% Plasma protein bound.
  • Onset of Action: 3-15 minutes.
  • Elimination Half-Life: 1.5-3 hours.

Society and Culture

Cost

Midazolam is available as a generic medication.

Availability

Midazolam is available in the United States as a syrup or as an injectable solution.

Dormicum brand midazolam is marketed by Roche as white, oval, 7.5-mg tablets in boxes of two or three blister strips of 10 tablets, and as blue, oval, 15-mg tablets in boxes of two (Dormonid 3x) blister strips of 10 tablets. The tablets are imprinted with “Roche” on one side and the dose of the tablet on the other side. Dormicum is also available as 1-, 3-, and 10-ml ampoules at a concentration of 5 mg/ml. Another manufacturer, Novell Pharmaceutical Laboratories, makes it available as Miloz in 3- and 5-ml ampoules. Midazolam is the only water-soluble benzodiazepine available. Another maker is Roxane Laboratories; the product in an oral solution, Midazolam HCl Syrup, 2 mg/ml clear, in a red to purplish-red syrup, cherry in flavour. It becomes soluble when the injectable solution is buffered to a pH of 2.9-3.7. Midazolam is also available in liquid form. It can be administered intramuscularly, intravenously, intrathecally, intranasally, buccally, or orally.

Legal Status

In the Netherlands, midazolam is a List II drug of the Opium Law. Midazolam is a Schedule IV drug under the Convention on Psychotropic Substances. In the United Kingdom, midazolam is a Schedule 3/Class C controlled drug. In the United States, midazolam (DEA number 2884) is on the Schedule IV list of the Controlled Substances Act as a non-narcotic agent with low potential for abuse.

Marketing Authorisation

In 2011, the European Medicines Agency (EMA) granted a marketing authorisation for a buccal application form of midazolam, sold under the trade name Buccolam. Buccolam was approved for the treatment of prolonged, acute, convulsive seizures in people from three months to less than 18 years of age. This was the first application of a paediatric-use marketing authorisation.

Use in Executions

The drug has been introduced for use in executions by lethal injection in certain jurisdictions in the United States in combination with other drugs. It was introduced to replace pentobarbital after the latter’s manufacturer disallowed that drug’s use for executions. Midazolam acts as a sedative but will fail to render the condemned prisoner unconscious, at which time vecuronium bromide and potassium chloride are administered, stopping the prisoner’s breathing and heart, respectively. Due to the fact that the condemned prisoner is not unconscious but merely sedated, two very different things, those following two drugs can cause extreme pain and panic in the soon to die prisoner.

Midazolam has been used as part of a three-drug cocktail, with vecuronium bromide and potassium chloride in Florida and Oklahoma prisons. Midazolam has also been used along with hydromorphone in a two-drug protocol in Ohio and Arizona.

The usage of midazolam in executions became controversial after condemned inmate Clayton Lockett apparently regained consciousness and started speaking midway through his 2014 execution when the state of Oklahoma attempted to execute him with an untested three-drug lethal injection combination using 100 mg of midazolam. Prison officials reportedly discussed taking him to a hospital before he was pronounced dead of a heart attack 40 minutes after the execution began. An observing doctor stated that Lockett’s vein had ruptured. It is not clear whether his death was caused by one or more of the drugs or to a problem in the administration procedure, nor is it clear what quantities of vecuronium bromide and potassium chloride were released to his system before the execution was cancelled.

Notable Incidents

The state of Florida used midazolam to execute William Frederick Happ in October 2013.

The state of Ohio used midazolam in the execution of Dennis McGuire in January 2014; it took McGuire 24 minutes to die after the procedure started, and he gasped and appeared to be choking during that time, leading to questions about the dosing and timing of the drug administration, as well as the choice of drugs.

The execution of Ronald Bert Smith in the state of Alabama on 08 December 2016, “went awry soon after (midazolam) was administered” again putting the effectiveness of the drug in question.

In October 2016, the state of Ohio announced that it would resume executions in January 2017, using a formulation of midazolam, vecuronium bromide, and potassium chloride, but this was blocked by a Federal judge. On 26 July 2017, Ronald Phillips was executed with a three-drug cocktail including midazolam after the Supreme Court refused to grant a stay. Prior to this, the last execution in Ohio had been that of Dennis McGuire. Murderer Gary Otte’s lawyers unsuccessfully challenged his Ohio execution, arguing that midazolam might not protect him from serious pain when the other drugs are administered. He died without incident in about 14 minutes on 13 September 2017.

On 24 April 2017, the state of Arkansas carried out a double-execution of Jack Harold Jones, 52, and Marcel Williams, 46. The state of Arkansas attempted to execute eight people before its supply of midazolam expired on 30 April 2017. Two of them were granted a stay of execution, and another, Ledell T. Lee, 51, was executed on 20 April 2017.

On 28 October 2021, the state of Oklahoma carried out the execution of inmate John Marion Grant, 60, using midazolam as part of its three-drug cocktail hours after the US Supreme Court ruled to lift a stay of execution for Oklahoma death row inmates. The execution was the state’s first since 2015. Witnesses to the execution said that when the first drug, midazolam, began to flow at 4:09 pm, Grant started convulsing about two dozen times and vomited. Grant continued breathing, and a member of the execution team wiped the vomit off his face. At 4:15 pm., officials said Grant was unconscious, and he was pronounced dead at 4:21 pm.

Legal Challenges

In Glossip v. Gross, attorneys for three Oklahoma inmates argued that midazolam could not achieve the level of unconsciousness required for surgery, meaning severe pain and suffering was likely. They argued that midazolam was cruel and unusual punishment and thus contrary to the Eighth Amendment to the United States Constitution. In June 2015, the US Supreme Court ruled that they had failed to prove that midazolam was cruel and unusual when compared to known, available alternatives.

The state of Nevada is also known to use midazolam in execution procedures. In July 2018, one of the manufacturers accused state officials of obtaining the medication under false pretences. This incident was the first time a drug company successfully, though temporarily, halted an execution. A previous attempt in 2017, to halt an execution in the state of Arizona by another drug manufacturer was not successful.

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What is Difludiazepam?

Introduction

Difludiazepam is a benzodiazepine derivative which is the 2′,6′-difluoro derivative of fludiazepam.

Background

It was invented in the 1970s but was never marketed, and has been used as a research tool to help determine the shape and function of the GABAA receptors, at which it has an IC50 of 4.1nM.

Difludiazepam has subsequently been sold as a designer drug, and was first notified to the EMCDDA by Swedish authorities in 2017.

What is Bretazenil?

Introduction

Bretazenil (Ro16-6028) is an imidazopyrrolobenzodiazepine anxiolytic drug which is derived from the benzodiazepine family, and was invented in 1988.

It is most closely related in structure to the GABA antagonist flumazenil, although its effects are somewhat different. It is classified as a high-potency benzodiazepine due to its high affinity binding to benzodiazepine binding sites where it acts as a partial agonist. Its profile as a partial agonist and preclinical trial data suggests that it may have a reduced adverse effect profile. In particular bretazenil has been proposed to cause a less strong development of tolerance and withdrawal syndrome. Bretazenil differs from traditional 1,4-benzodiazepines by being a partial agonist and because it binds to α1, α2, α3, α4, α5 and α6 subunit containing GABAA receptor benzodiazepine receptor complexes. 1,4-benzodiazepines bind only to α1, α2, α3 and α5 GABAA benzodiazepine receptor complexes.

Brief History

Bretazenil was originally developed as an anti-anxiety drug and has been studied for its use as an anticonvulsant but has never commercialised. It is a partial agonist for GABAA receptors in the brain. David Nutt from the University of Bristol has suggested bretazenil as a possible base from which to make a better social drug, as it displays several of the positive effects of alcohol intoxication such as relaxation and sociability, but without the bad effects such as aggression, amnesia, nausea, loss of coordination, liver disease and brain damage. The effects of bretazenil can also be quickly reversed by the action of flumazenil, which is used as an antidote to benzodiazepine overdose, in contrast to alcohol for which there is no effective and reliable antidote.

Traditional benzodiazepines are associated with side effects such as drowsiness, physical dependence and abuse potential. It was hoped that bretazenil and other partial agonists would be an improvement on traditional benzodiazepines which are full agonists due to preclinical evidence that their side effect profile was less than that of full agonist benzodiazepines. For a variety of reasons however, bretazenil and other partial agonists such as pazinaclone and abecarnil were not clinically successful. However, research continues into other compounds with partial agonist and compounds which are selective for certain GABAA benzodiazepine receptor subtypes.

Tolerance and Dependence

In a study in rats, cross-tolerance between the benzodiazepine drug chlordiazepoxide and bretazenil has been demonstrated. In a primate study bretazenil was found to be able to replace the full agonist diazepam in diazepam dependent primates without precipitating withdrawal effects, demonstrating cross tolerance between bretazenil and benzodiazepine agonists, whereas other partial agonists precipitated a withdrawal syndrome. The differences are likely due to differences in intrinsic properties between different benzodiazepine partial agonists. Cross-tolerance has also been shown between bretazenil and full agonist benzodiazepines in rats. In rats tolerance is slower to develop to the anticonvulsant effects compared to the benzodiazepine site full agonist diazepam. However, tolerance developed to the anticonvulsant effects of bretazenil partial agonist more quickly than they developed to imidazenil.

Pharmacology

Bretazenil has a more broad spectrum of action than traditional benzodiazepines as it has been shown to have low affinity binding to α4 and α6 GABAA receptors in addition to acting on α1, α2, α3 and α5 subunits which traditional benzodiazepine drugs work on. The partial agonist imidazenil does not, however, act at these subunits. 0.5mg of bretazenil is approximately equivalent in its psychomotor-impairing effect to 10 mg of diazepam. Bretazenil produces marked sedative-hypnotic effects when taken alone and when combined with alcohol. This human study also indicates that bretazenil is possibly more sedative than diazepam. The reason is unknown, but the study suggests the possibility that a full-agonist metabolite may be generated in humans but not animals previously tested or else that there are significant differences in benzodiazepine receptor population in animals and humans.

In a study of monkeys bretazenil has been found to antagonize the effects of full agonist benzodiazepines. However, bretazenil has been found to enhance the effects of neurosteroids acting on the neurosteroid binding site of the GABAA receptor. Another study found that bretazenil acted as an antagonist provoking withdrawal symptoms in monkeys who were physically dependent on the full agonist benzodiazepine triazolam.

Partial agonists of benzodiazepine receptors have been proposed as a possible alternative to full agonists of the benzodiazepine site to overcome the problems of tolerance, dependence and withdrawal which limits the role of benzodiazepines in the treatment of anxiety, insomnia and epilepsy. Such adverse effects appear to be less problematic with bretazenil than full agonists. Bretazenil has also been found to have less abuse potential than benzodiazepine full agonists such as diazepam and alprazolam, however long-term use of bretazenil would still be expected to result in dependence and addiction.

Bretazenil alters the sleep EEG profile and causes a reduction in cortisol secretion and increases significantly the release of prolactin. Bretazenil has effective hypnotic properties but impairs cognitive ability in humans. Bretazenil causes a reduction in the number of movements between sleep stages and delays movement into REM sleep. At a dosage of 0.5 mg of bretazenil REM sleep is decreased and stage 2 sleep is lengthened.

What is Ramelteon?

Introduction

Ramelteon, sold under the brand name Rozerem among others, is a sleep agent medication that selectively binds to the MT1 and MT2 receptors in the suprachiasmatic nucleus (SCN), instead of binding to GABAA receptors, such as with drugs like zolpidem.

It appears to speed the onset of sleep and alter the total amount of sleep a person gets. It is approved by the US Food and Drug Administration (FDA) for long-term use.

Ramelteon does not show any appreciable binding to GABAA receptors, which are associated with anxiolytic, myorelaxant, and amnesic effects.

Brief History

Ramelteon was approved for use in the United States in July 2005.

Medical Uses

Ramelteon is approved in the United States for the treatment of insomnia characterised by difficulty with sleep onset.

A systematic review, published in 2014, concluded “ramelteon was found to be beneficial in preventing delirium in medically ill individuals when compared to placebo.”

Mechanism of Action

Ramelteon is a melatonin receptor agonist with both high affinity for melatonin MT1 and MT2 receptors and selectivity over the MT3 receptor. Ramelteon demonstrates full agonist activity in vitro in cells expressing human MT1 or MT2 receptors, and high selectivity for human MT1 and MT2 receptors compared to the MT3 receptor.

The activity of ramelteon at the MT1 and MT2 receptors is believed to contribute to its sleep-promoting properties, as these receptors, acted upon by endogenous melatonin, are thought to be involved in the maintenance of the circadian rhythm underlying the normal sleep-wake cycle. Ramelteon has no appreciable affinity for the GABA receptor complex or for receptors that bind neuropeptides, cytokines, serotonin, dopamine, noradrenaline, acetylcholine, and opioids. Ramelteon also does not interfere with the activity of a number of selected enzymes in a standard panel.

The major metabolite of ramelteon, M-II, is active and has approximately one tenth and one fifth the binding affinity of the parent molecule for the human MT1 and MT2 receptors, respectively, and is 17-25-fold less potent than ramelteon in in vitro functional assays. Although the potency of M-II at MT1 and MT2 receptors is lower than the parent drug, M-II circulates at higher concentrations than the parent producing 20-100-fold greater mean systemic exposure when compared to ramelteon. M-II has weak affinity for the serotonin 5-HT2B receptor, but no appreciable affinity for other receptors or enzymes. Similar to ramelteon, M-II does not interfere with the activity of a number of endogenous enzymes.

Adverse Effects

Ramelteon has not been shown to produce dependence and has shown no potential for abuse, and the withdrawal and rebound insomnia that is typical with GABA modulators is not present in ramelteon.

Six percent of ramelteon-treated patients in clinical trials discontinued due to an adverse event, compared with two percent in the placebo arms. The most frequent adverse events leading to discontinuation were somnolence, dizziness, nausea, fatigue, headache, and insomnia. The US official Prescribing Information warns of rare cases of anaphylactic reactions, abnormal thinking, worsening of depression or suicidal thinking in patients with pre-existing depression, and decreased testosterone and increased prolactin levels. It also notes that ramelteon is not recommended for use in patients with severe sleep apnoea.

In mice treated with ramelteon for two years, increases in liver and testicular tumours were observed, but only at doses at least 20 times greater than the recommended human dose on a milligram/kilogram basis.

Drug Interactions

Ramelteon has been evaluated for potential drug interactions with the following medications and showed no significant effects: omeprazole, theophylline, dextromethorphan, and midazolam, digoxin and warfarin. There were no clinically meaningful effects when ramelteon was co-administered with any of these drugs.

A drug interaction study showed that there were no clinically meaningful effects or an increase in adverse events when ramelteon and the SSRI Prozac (fluoxetine) were co-administered. When co-administered with ramelteon, fluvoxamine (strong CYP1A2 inhibitor) increased AUC approximately 190-fold, and the Cmax increased approximately 70-fold, compared to ramelteon administered alone. Ramelteon and fluvoxamine should not be co-administered.

Ramelteon has significant drug-drug interaction with the following drugs: amiodarone, ciprofloxacin, fluvoxamine, ticlopidine.

Ramelteon should be administered with caution in patients taking other CYP1A2 inhibitors, strong CYP3A4 inhibitors such as ketoconazole, and strong CYP2C9 inhibitors such as fluconazole.

Efficacy may be reduced when ramelteon is used in combination with potent CYP enzyme inducers such as rifampin, since ramelteon concentrations may be decreased.

What is a GABA Receptor?

Introduction

The GABA receptors are a class of receptors that respond to the neurotransmitter gamma-aminobutyric acid (GABA), the chief inhibitory compound in the mature vertebrate central nervous system.

There are two classes of GABA receptors: GABAA and GABAB. GABAA receptors are ligand-gated ion channels (also known as ionotropic receptors); whereas GABAB receptors are G protein-coupled receptors, also called metabotropic receptors.

Ligand-Gated Ion Channels

Ionotropic GABA receptors (iGABARs) are ligand-gated ion channel of the GABA receptors class which are activated by gamma-aminobutyric acid (GABA), and include:

  • GABAA receptors.
  • GABAA-ρ receptors.

The GABAB receptor, a G protein-coupled receptor, is the only metabotropic GABA receptor and its mechanism of action differs significantly from the ionotropic receptors. Functionally, in mature organisms, activation of these receptors typically results in neural inhibition, primarily via the influx of chloride ions, although exceptions to this general principle exist, such as during early development. Structurally, iGABARs are pentameric transmembrane ion channels, meaning they are made up of five subunits. Since there are several classes of subunits and a variety of genes encoding many members of these classes, a wide variety of structurally, and therefore functionally, distinct channels of iGABARs is observed.

GABAA Receptor

It has long been recognised that the fast response of neurons to GABA that is stimulated by bicuculline and picrotoxin is due to direct activation of an anion channel. This channel was subsequently termed the GABAA receptor. Fast-responding GABA receptors are members of a family of Cys-loop ligand-gated ion channels. Members of this superfamily, which includes nicotinic acetylcholine receptors, GABAA receptors, glycine and 5-HT3 receptors, possess a characteristic loop formed by a disulfide bond between two cysteine residues.

In ionotropic GABAA receptors, binding of GABA molecules to their binding sites in the extracellular part of the receptor triggers opening of a chloride ion-selective pore. The increased chloride conductance drives the membrane potential towards the reversal potential of the Cl¯ ion which is about -75 mV in neurons, inhibiting the firing of new action potentials. This mechanism is responsible for the sedative effects of GABAA allosteric agonists. In addition, activation of GABA receptors lead to the so-called shunting inhibition, which reduces the excitability of the cell independent of the changes in membrane potential.

There have been numerous reports of excitatory GABAA receptors. According to the excitatory GABA theory, this phenomenon is due to increased intracellular concentration of Cl¯ ions either during development of the nervous system or in certain cell populations. After this period of development, a chloride pump is upregulated and inserted into the cell membrane, pumping Cl− ions into the extracellular space of the tissue. Further openings via GABA binding to the receptor then produce inhibitory responses. Over-excitation of this receptor induces receptor remodelling and the eventual invagination of the GABA receptor. As a result, further GABA binding becomes inhibited and inhibitory postsynaptic potentials are no longer relevant.

However, the excitatory GABA theory has been questioned as potentially being an artefact of experimental conditions, with most data acquired in in-vitro brain slice experiments susceptible to un-physiological milieu such as deficient energy metabolism and neuronal damage. The controversy arose when a number of studies have shown that GABA in neonatal brain slices becomes inhibitory if glucose in perfusate is supplemented with ketone bodies, pyruvate, or lactate, or that the excitatory GABA was an artefact of neuronal damage. Subsequent studies from originators and proponents of the excitatory GABA theory have questioned these results, but the truth remained elusive until the real effects of GABA could be reliably elucidated in intact living brain. Since then, using technology such as in-vivo electrophysiology/imaging and optogenetics, two in-vivo studies have reported the effect of GABA on neonatal brain, and both have shown that GABA is indeed overall inhibitory, with its activation in the developing rodent brain not resulting in network activation, and instead leading to a decrease of activity.

GABA receptors influence neural function by coordinating with glutamatergic processes.

GABAA-ρ Receptor

A subclass of ionotropic GABA receptors, insensitive to typical allosteric modulators of GABAA receptor channels such as benzodiazepines and barbiturates, was designated GABAС receptor. Native responses of the GABAC receptor type occur in retinal bipolar or horizontal cells across vertebrate species.

GABAС receptors are exclusively composed of ρ (rho) subunits that are related to GABAA receptor subunits. Although the term “GABAС receptor” is frequently used, GABAС may be viewed as a variant within the GABAA receptor family. Others have argued that the differences between GABAС and GABAA receptors are large enough to justify maintaining the distinction between these two subclasses of GABA receptors. However, since GABAС receptors are closely related in sequence, structure, and function to GABAA receptors and since other GABAA receptors besides those containing ρ subunits appear to exhibit GABAС pharmacology, the Nomenclature Committee of the IUPHAR has recommended that the GABAС term no longer be used and these ρ receptors should be designated as the ρ subfamily of the GABAA receptors (GABAA-ρ).

G Protein-Coupled Receptors

GABAB Receptor

A subclass of ionotropic GABA receptors, insensitive to typical allosteric modulators of GABAA receptor channels such as benzodiazepines and barbiturates, was designated GABAС receptor. Native responses of the GABAC receptor type occur in retinal bipolar or horizontal cells across vertebrate species.

GABAС receptors are exclusively composed of ρ (rho) subunits that are related to GABAA receptor subunits. Although the term “GABAС receptor” is frequently used, GABAС may be viewed as a variant within the GABAA receptor family. Others have argued that the differences between GABAС and GABAA receptors are large enough to justify maintaining the distinction between these two subclasses of GABA receptors. However, since GABAС receptors are closely related in sequence, structure, and function to GABAA receptors and since other GABAA receptors besides those containing ρ subunits appear to exhibit GABAС pharmacology, the Nomenclature Committee of the IUPHAR has recommended that the GABAС term no longer be used and these ρ receptors should be designated as the ρ subfamily of the GABAA receptors (GABAA-ρ).

GABA Receptor Gene Polymorphisms

Two separate genes on two chromosomes control GABA synthesis – glutamate decarboxylase and alpha-ketoglutarate decarboxylase genes – though not much research has been done to explain this polygenic phenomenon. GABA receptor genes have been studied more in depth, and many have hypothesized about the deleterious effects of polymorphisms in these receptor genes. The most common single nucleotide polymorphisms (SNPs) occurring in GABA receptor genes rho 1, 2, and 3 (GABBR1, GABBR2, and GABBR3) have been more recently explored in literature, in addition to the potential effects of these polymorphisms. However, some research has demonstrated that there is evidence that these polymorphisms caused by single base pair variations may be harmful.

It was discovered that the minor allele of a single nucleotide polymorphism at GABBR1 known as rs1186902 is significantly associated with a later age of onset for migraines, but for the other SNPs, no differences were discovered between genetic and allelic variations in the control vs. migraine participants. Similarly, in a study examining SNPs in rho 1, 2, and 3, and their implication in essential tremor, a nervous system disorder, it was discovered that there were no differences in the frequencies of the allelic variants of polymorphisms for control vs. essential tremor participants. On the other hand, research examining the effect of SNPs in participants with restless leg syndrome found an “association between GABRR3rs832032 polymorphism and the risk for RLS, and a modifier effect of GABRA4 rs2229940 on the age of onset of RLS” – the latter of which is a modifier gene polymorphism. The most common GABA receptor SNPs do not correlate with deleterious health effects in many cases, but do in a few.

One significant example of a deleterious mutation is the major association between several GABA receptor gene polymorphisms and schizophrenia. Because GABA is integral to the release of inhibitory neurotransmitters which produce a calming effect and play a role in reducing anxiety, stress, and fear, it is not surprising that polymorphisms in these genes result in more consequences relating to mental health than to physical health. Of an analysis on 19 SNPs on various GABA receptor genes, five SNPs in the GABBR2 group were found to be significantly associated with schizophrenia, which produce the unexpected haplotype frequencies not found in the studies mentioned previously.

Several studies have verified association between alcohol use disorder and the rs279858 polymorphism on the GABRA2 gene e, and higher negative alcohol effects scores for individuals who were homozygous at six SNPs. Furthermore, a study examining polymorphisms in the GABA receptor beta 2 subunit gene found an association with schizophrenia and bipolar disorder, and examined three SNPs and their effects on disease frequency and treatment dosage. A major finding of this study was that functional psychosis should be conceptualised as a scale of phenotypes rather than distinct categories.

What is Gamma-Aminobutyric Acid?

Introduction

gamma-Aminobutyric acid (or γ-aminobutyric acid or GABA), is the chief inhibitory neurotransmitter in the developmentally mature mammalian central nervous system. Its principal role is reducing neuronal excitability throughout the nervous system.

GABA is sold as a dietary supplement in many countries. It has been traditionally thought that exogenous GABA (i.e. taken as a supplement) does not cross the blood-brain barrier, however data obtained from more current research indicates that it may be possible.

Brief History

In 1883, GABA was first synthesized, and it was first known only as a plant and microbe metabolic product.

In 1950, GABA was discovered as an integral part of the mammalian central nervous system.

In 1959, it was shown that at an inhibitory synapse on crayfish muscle fibres GABA acts like stimulation of the inhibitory nerve. Both inhibition by nerve stimulation and by applied GABA are blocked by picrotoxin.

Function

Neurotransmitter

Refer to GABA Receptor.

Two general classes of GABA receptor are known:

  • GABAA in which the receptor is part of a ligand-gated ion channel complex; and
  • GABAB metabotropic receptors, which are G protein-coupled receptors that open or close ion channels via intermediaries (G proteins).

Neurons that produce GABA as their output are called GABAergic neurons, and have chiefly inhibitory action at receptors in the adult vertebrate. Medium spiny cells are a typical example of inhibitory central nervous system GABAergic cells. In contrast, GABA exhibits both excitatory and inhibitory actions in insects, mediating muscle activation at synapses between nerves and muscle cells, and also the stimulation of certain glands. In mammals, some GABAergic neurons, such as chandelier cells, are also able to excite their glutamatergic counterparts.

Release, Reuptake, and Metabolism Cycle of GABA.

GABAA receptors are ligand-activated chloride channels: when activated by GABA, they allow the flow of chloride ions across the membrane of the cell. Whether this chloride flow is depolarising (makes the voltage across the cell’s membrane less negative), shunting (has no effect on the cell’s membrane potential), or inhibitory/hyperpolarising (makes the cell’s membrane more negative) depends on the direction of the flow of chloride. When net chloride flows out of the cell, GABA is depolarising; when chloride flows into the cell, GABA is inhibitory or hyperpolarising. When the net flow of chloride is close to zero, the action of GABA is shunting. Shunting inhibition has no direct effect on the membrane potential of the cell; however, it reduces the effect of any coincident synaptic input by reducing the electrical resistance of the cell’s membrane. Shunting inhibition can “override” the excitatory effect of depolarising GABA, resulting in overall inhibition even if the membrane potential becomes less negative. It was thought that a developmental switch in the molecular machinery controlling the concentration of chloride inside the cell changes the functional role of GABA between neonatal and adult stages. As the brain develops into adulthood, GABA’s role changes from excitatory to inhibitory.

Brain Development

While GABA is an inhibitory transmitter in the mature brain, its actions were thought to be primarily excitatory in the developing brain. The gradient of chloride was reported to be reversed in immature neurons, with its reversal potential higher than the resting membrane potential of the cell; activation of a GABA-A receptor thus leads to efflux of Cl− ions from the cell (that is, a depolarising current). The differential gradient of chloride in immature neurons was shown to be primarily due to the higher concentration of NKCC1 co-transporters relative to KCC2 co-transporters in immature cells. GABAergic interneurons mature faster in the hippocampus and the GABA signalling machinery appears earlier than glutamatergic transmission. Thus, GABA is considered the major excitatory neurotransmitter in many regions of the brain before the maturation of glutamatergic synapses.

In the developmental stages preceding the formation of synaptic contacts, GABA is synthesized by neurons and acts both as an autocrine (acting on the same cell) and paracrine (acting on nearby cells) signalling mediator. The ganglionic eminences also contribute greatly to building up the GABAergic cortical cell population.

GABA regulates the proliferation of neural progenitor cells, the migration and differentiation the elongation of neurites and the formation of synapses.

GABA also regulates the growth of embryonic and neural stem cells. GABA can influence the development of neural progenitor cells via brain-derived neurotrophic factor (BDNF) expression. GABA activates the GABAA receptor, causing cell cycle arrest in the S-phase, limiting growth.

Beyond the Nervous System

Besides the nervous system, GABA is also produced at relatively high levels in the insulin-producing β-cells of the pancreas. The β-cells secrete GABA along with insulin and the GABA binds to GABA receptors on the neighbouring islet α-cells and inhibits them from secreting glucagon (which would counteract insulin’s effects).

GABA can promote the replication and survival of β-cells and also promote the conversion of α-cells to β-cells, which may lead to new treatments for diabetes.

Alongside GABAergic mechanisms, GABA has also been detected in other peripheral tissues including intestines, stomach, Fallopian tubes, uterus, ovaries, testes, kidneys, urinary bladder, the lungs and liver, albeit at much lower levels than in neurons or β-cells.

Experiments on mice have shown that hypothyroidism induced by fluoride poisoning can be halted by administering GABA. The test also found that the thyroid recovered naturally without further assistance after the Fluoride had been expelled by the GABA.

Immune cells express receptors for GABA and administration of GABA can suppress inflammatory immune responses and promote “regulatory” immune responses, such that GABA administration has been shown to inhibit autoimmune diseases in several animal models.

In 2018, GABA has shown to regulate secretion of a greater number of cytokines. In plasma of T1D patients, levels of 26 cytokines are increased and of those, 16 are inhibited by GABA in the cell assays.

In 2007, an excitatory GABAergic system was described in the airway epithelium. The system is activated by exposure to allergens and may participate in the mechanisms of asthma. GABAergic systems have also been found in the testis and in the eye lens.

GABA occurs in plants.

Structure and Conformation

GABA is found mostly as a zwitterion (i.e. with the carboxyl group deprotonated and the amino group protonated). Its conformation depends on its environment. In the gas phase, a highly folded conformation is strongly favoured due to the electrostatic attraction between the two functional groups. The stabilisation is about 50 kcal/mol, according to quantum chemistry calculations. In the solid state, an extended conformation is found, with a trans conformation at the amino end and a gauche conformation at the carboxyl end. This is due to the packing interactions with the neighbouring molecules. In solution, five different conformations, some folded and some extended, are found as a result of solvation effects. The conformational flexibility of GABA is important for its biological function, as it has been found to bind to different receptors with different conformations. Many GABA analogues with pharmaceutical applications have more rigid structures in order to control the binding better.

Biosynthesis

GABA is primarily synthesized from glutamate via the enzyme glutamate decarboxylase (GAD) with pyridoxal phosphate (the active form of vitamin B6) as a cofactor. This process converts glutamate (the principal excitatory neurotransmitter) into GABA (the principal inhibitory neurotransmitter).

GABA can also be synthesized from putrescine by diamine oxidase and aldehyde dehydrogenase.

Traditionally it was thought that exogenous GABA did not penetrate the blood–brain barrier, however more current research indicates that it may be possible, or that exogenous GABA (i.e. in the form of nutritional supplements) could exert GABAergic effects on the enteric nervous system which in turn stimulate endogenous GABA production. The direct involvement of GABA in the glutamate-glutamine cycle makes the question of whether GABA can penetrate the blood-brain barrier somewhat misleading, because both glutamate and glutamine can freely cross the barrier and convert to GABA within the brain.

Metabolism

GABA transaminase enzymes catalyse the conversion of 4-aminobutanoic acid (GABA) and 2-oxoglutarate (α-ketoglutarate) into succinic semialdehyde and glutamate. Succinic semialdehyde is then oxidized into succinic acid by succinic semialdehyde dehydrogenase and as such enters the citric acid cycle as a usable source of energy.

Pharmacology

Drugs that act as allosteric modulators of GABA receptors (known as GABA analogues or GABAergic drugs), or increase the available amount of GABA, typically have relaxing, anti-anxiety, and anti-convulsive effects. Many of the substances below are known to cause anterograde amnesia and retrograde amnesia.

In general, GABA does not cross the blood-brain barrier, although certain areas of the brain that have no effective blood–brain barrier, such as the periventricular nucleus, can be reached by drugs such as systemically injected GABA. At least one study suggests that orally administered GABA increases the amount of human growth hormone (HGH). GABA directly injected to the brain has been reported to have both stimulatory and inhibitory effects on the production of growth hormone, depending on the physiology of the individual. Certain pro-drugs of GABA (e.g. picamilon) have been developed to permeate the blood-brain barrier, then separate into GABA and the carrier molecule once inside the brain. Prodrugs allow for a direct increase of GABA levels throughout all areas of the brain, in a manner following the distribution pattern of the pro-drug prior to metabolism.

GABA enhanced the catabolism of serotonin into N-acetylserotonin (the precursor of melatonin) in rats. It is thus suspected that GABA is involved in the synthesis of melatonin and thus might exert regulatory effects on sleep and reproductive functions.

Chemistry

Although in chemical terms, GABA is an amino acid (as it has both a primary amine and a carboxylic acid functional group), it is rarely referred to as such in the professional, scientific, or medical community. By convention the term “amino acid”, when used without a qualifier, refers specifically to an alpha amino acid. GABA is not an alpha amino acid, meaning the amino group is not attached to the alpha carbon. Nor is it incorporated into proteins as are many alpha-amino acids.

GABAergic Drugs

GABAA receptor ligands are shown in the following table.

Activity at GABAALigand
Orthosteric AgonistMuscimol, GABA, gaboxadol (THIP), isoguvacine, progabide, piperidine-4-sulfonic acid (partial agonist).
Positive allosteric modulatorsBarbiturates, benzodiazepines, neuroactive steroids, niacin/niacinamide, nonbenzodiazepines (i.e. z-drugs, e.g. zolpidem, eszopiclone), etomidate, etaqualone, alcohol (ethanol), theanine, methaqualone, propofol, stiripentol, and anaesthetics (including volatile anaesthetics), glutethimide.
Orthosteric (competitive) AntagonistBicuculline, gabazine, thujone, and flumazenil.
Uncompetitive antagonist (e.g. channel blocker)Picrotoxin, and cicutoxin.
Negative allosteric modulatorsNeuroactive steroids (Pregnenolone sulfate), furosemide, oenanthotoxin, and amentoflavone.

Additionally, carisoprodol is an enhancer of GABAA activity. Ro15-4513 is a reducer of GABAA activity

GABAergic pro-drugs include chloral hydrate, which is metabolised to trichloroethanol, which then acts via the GABAA receptor.

Skullcap and valerian are plants containing GABAergic substances[citation needed]. Furthermore, the plant kava contains GABAergic compounds, including kavain, dihydrokavain, methysticin, dihydromethysticin and yangonin.

Other GABAergic modulators include:

  • GABAB receptor ligands.
    • Agonists: baclofen, propofol, GHB, and phenibut.
    • Antagonists: phaclofen and saclofen.
  • GABA reuptake inhibitors: deramciclane, hyperforin, and tiagabine.
  • GABA transaminase inhibitors: gabaculine, phenelzine, valproate, vigabatrin, and lemon balm (Melissa officinalis).
  • GABA analogues: pregabalin, gabapentin, picamilon, and progabide.

In Plants

GABA is also found in plants. It is the most abundant amino acid in the apoplast of tomatoes. Evidence also suggests a role in cell signalling in plants.