Presently, Adderall is the 27th most prescribed medication in the United States, with over 24 million prescriptions in 2020. Given its accelerating use, including non-medically as a “study drug,” an in-depth understanding of how this drug exerts its effects on cognition and behavior is warranted.
To this end, rodent models have been tremendously helpful in uncovering the neurobiological mechanisms of Adderall use and its behavioral correlates. In this article, we will look at some research findings and experiments within the Adderall literature, including the methods used by the researchers.
What is Adderall?
Adderall: A medical drug
Adderall is a powerful central nervous stimulant composed of a racemic mixture of amphetamine salts.
- 75% D-amphetamine (dextroamphetamine):, the right-handed enantiomer of amphetamine that has a more pronounced stimulating effect on the central nervous system.
- 25% L-amphetamine (levoamphetamine): the left-handed enantiomer of amphetamine that has more potent peripheral effects due to its noradrenergic selectivity.
Its active ingredient is amphetamine, which is classified by the Drug Enforcement Administration (DEA) as a Schedule II drug. This implies it has a high potential for dependence and abuse but also carries medical uses, as we will discuss further below.
Regardless of its role in medicine, Adderall’s side effects can be serious, including: appetite suppression, weight loss, abdominal pain headaches, sleep disturbances, dizziness, nausea, and tremor. In rare cases, Adderall can even cause hypertension, mania, hallucinations, aggression, and dysphoria. Additionally, being a powerful central nervous system stimulant affecting the peripheral system as well, Adderall is contraindicated in users with underlying cardiovascular conditions or a history of psychosis.
Historical Origins of Adderall
Amphetamine was originally derived from ephedrine, an extract of the Ephedra plant that is used for allergies, asthma, and colds. It’s first synthesis dates back to 1887, but its stimulant effects were not uncovered until 1927.
In World War II, amphetamine and dextroamphetamine (under the brand names Benzedrine and Dexedrine) were widely used by both the Allied and Axis forces to combat fatigue and boost alertness and performance. By the 1950s, Benzedrine became popular among general Americans, where it was extensively used for weight loss, cognitive enhancement, and to treat Parkinson’s disease.
The first amphetamine mixed salts came to the market as Obetrol. This was indicated to treat obesity by the American pharmaceutical company Rexar. Although Obetrol was unapproved by the FDA at the time, Rexar kept selling amphetamine for weight loss into the 1990s. In 1993, Rexar was bought by Richwood Pharmaceuticals, who rebranded Obetrol as Adderall, a marketing-infused word derived from “A.D.D. for All”.
By 1996, Richwood pharmaceuticals (who later merged with Shire Pharmaceuticals) introduced the first instant-release Adderall tablet. Following its major success, this was later followed by the extended-release formulation in 2001. Today, the ADHD drug industry makes more than 13 billion dollars annually and continues to climb with the accelerating use of these drugs among children and adults alike.
Adderall’s Therapeutic Properties
Adderall is prescribed primarily for Attention Deficit Hyperactivity Disorder (ADHD) and Attention Deficit Disorder (ADD), but also for narcolepsy. In some cases, it’s prescribed off-label for atypical anxiety and depression.
To target ADHD symptoms, Adderall is prescribed in two different forms: instant release (IR) or extended-release (XR). The IR formulation has a half-life (the time it takes for the body to excrete half of the drug) of approximately 9 hours, whereas the XR formulation has a half-life of 10-13 hours. The XR formulation is usually taken in the morning and nearly doubles the duration of the effect. In terms of dosing, a 20mg dose of XR once per day is equivalent to a 10mg IR dose taken twice per day. Thus, these two different forms of Adderall are able to be prescribed according to the patient’s needs and lifestyle.
How Does Adderall Work?
Adderall exerts its effects through its interaction with monoamine transporters, inducing neurotransmitter release from neuron terminals. This includes mainly dopamine, but also serotonin and norepinephrine.
Amphetamine enters neurons by binding and interacting with presynaptic monoamine reuptake transporters and receptors:
- Trace amine-associated receptor 1 (TAAR1): A G protein-coupled receptor expressed within the presynaptic terminal of monoamine neurons that are activated by amphetamine.
- Vesicular monoamine transporter 2 (VMAT2): An integral membrane protein that transports monoamines from cellular cytosol to synaptic vesicles.
Amphetamine’s action on VMAT2 results in higher levels of neurotransmitters within the neuron. When amphetamine activates TAAR1, the receptor induces the neuron’s cell membrane-bound monoamine transporters to either stop transporting monoamines altogether or transport monoamines out of the neuron. Higher cytoplasmic concentrations of the neurotransmitters, coupled with amphetamine’s action on VMAT2, cause monoamine transporters to reverse its direction of transport. The reversed membrane transporter will, therefore, push dopamine, norepinephrine, and serotonin out of the neuron’s intracellular fluid and into the synaptic cleft. This leads to an increased release of neurotransmitters and, consequently, higher concentrations within the synaptic cleft.
|Adderall’s Effects on TAAR1||Adderall’s Effects on VMAT2|
|● TAAR1 activation causes an inhibition of dopamine uptake by Dopamine Active Transporter (DAT).|
● Enhanced TAAR1 the signaling increases production of the second messenger cyclic AMP (cAMP).
● TAAR1 activation enhances protein kinase signaling, resulting in DAT phosphorylation and reverses the direction of transport.
|● Binds to distinct sites on VMAT2 and inhibits its function.|
● Inhibition of VMAT2 releases monoamine neurotransmitters from vesicles into the intracellular fluid and prevents the reuptake of monoamine neurotransmitters.
Caption: General summary of how Adderall affects neuronal receptors and reuptake transporters.[5-6]
Being an amphetamine salt mixture, the D-amphetamine and L-amphetamine isomers exhibit different pharmacodynamics from each other. D-amphetamine has stronger effects on the central nervous system, whereas L-amphetamine has greater peripheral effects as well as a longer half-life. Additionally, L-amphetamine alters the dopamine release kinetics, resulting in a faster dopamine rise time and shorter decay times.
The large majority of animal studies utilize dextroamphetamine (Dexedrine). We will be focusing on the studies that use the racemic mixture D,L-amphetamine in order to best capture the effects of Adderall on rodent behavior.
The Cognitive Effects of Adderall at a Glance
Overall, the enhanced signaling and release of these neurotransmitters leads to wide-ranging effects on the central nervous system. In humans, enhanced dopaminergic and serotonergic signaling exert a wide array of effects on behavior and mood, enhancing focus, energy, attention, wakefulness, and producing euphoria in higher doses.
Through enhanced epinephrine signaling, Adderall activates the sympathetic nervous system. This leads to increased alertness and a peripheral flight-or-flight response characterized by increased heart rate, blood pressure, slowed digestion, and pupil dilation.
Elevated levels of dopamine within the mesolimbic dopamine reward pathway, especially within the nucleus accumbens, underlie its strong psychomotor rewarding and addictive properties.
With repeated dosing of Adderall, especially at non-therapeutic (recreational) doses, desensitization of the reward pathway begins to occur, causing the user to require more and more of the drug for the same effects. Given that Adderall is increasingly being used non-medically as a “study drug” to increase performance in work or school, it’s important to underscore Adderall’s abuse potential, especially when used chronically and in high doses.
Armed with an understanding of Adderall’s mechanism of action and general effect profile, let’s take a closer look at how Adderall impacts rodent behavior.
Adderall’s Effects on ADHD
ADHD is behaviorally characterized by impulsivity, inattention, and hyperactivity. These symptoms are thought to result from the dysfunctional release of dopamine, though research is still mixed as to whether ADHD represents a hyperdopaminergic or hypodopaminergic state.
However, imaging studies of individuals with ADHD have consistently shown abnormalities in brain regions dealing with motor coordination and executive control. Specifically, they show smaller sizes of dopaminergic regions, including the striatum, basal ganglia, and prefrontal cortex.
Interestingly, one of the behavioral hallmarks of ADHD is that amphetamine treatment produces paradoxical calming effects, helping to reduce symptoms of hyperactivity and impulsivity. Below, we’ll see that this effect is also recapitulated in rodent models of ADHD.
The 6-OHDA Murine Model of ADHD
The 6-hydroxydopamine-lesioned (6-OHDA) murine model is a widely used animal model of ADHD. Through the neonatal lesioning of nigrostriatal dopaminergic pathways, this model produces dopamine-deficient mice that exhibit typical ADHD symptoms. This includes hyperactivity, deficits in behavioral inhibition, and higher instances of impulsivity.
A study by Avale et al. investigated the behavioral effects of D,L-amphetamine on 6-OHDA mice. However, Avale and colleagues first recapitulated many behavioral features of ADHD in this mouse model at postnatal day 24 (weaning age). Compared to controls, the 6-OHDA mice demonstrated hyperlocomotion in an open field arena. This was seen as a higher number of movement initiations and increased vertical exploratory activity (rearing). These effects were noted for several weeks but were found to wane near puberty, around 10 weeks postnatal.
The 6-OHDA mice also demonstrated deficits in behavioral inhibition in the open field arena. While control mice avoided the central part of the maze (an open, unsafe area), the 6-OHDA mice spent more time in the unprotected parts of this novel environment compared to controls, indicating less behavioral inhibition.
Amphetamine Salts Produce Hyperactivity and Decreased Behavioral Inhibition in 6-OHDA Mice
Avale and colleagues then administered intraperitoneal injections of D,L-amphetamine at 4mg/kg to a group of 6-OHDA mice and controls. While the control mice showed increased hyperactivity in the open field arena, the 6-OHDA mice demonstrated a paradoxical hypolocomotor effect, seen as significant reductions in horizontal exploratory activity. These findings mimic key features of ADHD in humans, namely, the paradoxical effect of amphetamine treatment as discussed earlier.
Dopamine Receptor D4 Mediates Hyperactivity and Behavioral Inhibition
The researchers then honed in on specific ADHD candidate genes that may be responsible for the ADHD symptomatology, especially to understand the manifestation of the paradoxical hypolocomotion effect in ADHD mice given Adderall. To this end, they bred mice lacking D4R receptors (G-protein coupled receptors that are known to play a major role in mediating dopamine-related functions), especially within prefrontal cortical circuits. The rationale for this gene choice was related to the fact that specific polymorphisms of this gene are known to occur at high rates in individuals with ADHD, a disorder which is also known for its high heritability (50-90% according to twin studies).
Interestingly, the researchers found that 6-OHDA mice lacking the D4R receptors did not develop hyperactivity in the open field arena like their wildtype counterparts, nor did they exhibit high amounts of behavioral disinhibition. The influence of D4 receptors on hyperactivity was further validated when they injected the non-mutant, normal 6-OHDA mice with PNU-101387G, a D4 receptor antagonist. This antagonist reduced locomotor activity levels but had no effect on amphetamine-induced hyperactivity.
When given D,L-amphetamine injections at 4mg/kg, the D4R knockout 6-OHDA mice showed less spontaneous locomotor activity in the open arena, similar to what was seen in the wildtype mice. Thus, D,L-amphetamine administration was found to decrease locomotor activity in 6-OHDA mice and increase locomotor activity in non-lesioned mice, regardless of genotype in both cases.
|Mouse Strain||Behavioral Profile|
|Normal Controls||● Normal baseline activity in the open field arena.|
● Behavioral inhibition, seen as less time spent in the central part of the open field arena
|● Enhanced locomotion in the open field arena, seen as a nearly two-fold increase in horizontal activity.|
|6-OHDA Mice||● A two-fold increase in spontaneous hyperlocomotion (vertical and horizontal activity) in the open field arena compared to normal controls|
● Decreased behavioral inhibition, reflected as more time spent in the central part of the open field arena (47% of total distance traveled in central zone vs 28% in controls)
|● Paradoxical hypolocomotion in open field arena, seen as less horizontal activity in the arena compared to normal controls|
|DRD4 KO Control Mice||● Similar activity scores in the open field arena as wildtype controls|
● Behavioral inhibition in the central part of the open field arena, similar to wildtype controls
|DRD4 KO Control MIce|
|● Enhanced locomotion in open field arena, similar to wildtype controls|
|DRD4 KO 6-OHDA Mice||● No observed hyperactivity in open field arena, identical activity scores as control mice|
● Increased behavioral inhibition, seen as less time spent in the central part of the open field arena
|DRD4 KO 6-OHDA Mice + Adderall||● Paradoxical hypolocomotion in open field arena, seen as decreased locomotion scores by 50% compared to DRD4 KO control mice|
Caption: Behavioral profiles of wildtype and mutant mice with and without 6-OHDA lesions. DRD4 activation was found to be necessary for the behavioral disinhibition and hyperactivity observed in the 6-OHDA phenotype. However, D,L-amphetamine’s paradoxical hypolocomotion effect in the open field arena is independent of DRD4.
The researchers conclude that the paradoxical response to D,L-amphetamine in this ADHD-like model is independent of dopamine receptor D4. Instead, the observed effects of D,L-amphetamine are probably related to the alteration of other dopamine receptors or the interaction with neurotransmitters other than dopamine. Though, with regards to dopamine, D1 and D2 dopamine receptors are likely candidates given how tightly linked they are to motor execution and planning.
Adderall’s Effects on Anxiety
Chronic Adderall treatment can create nervousness, anxiety, and social withdrawal in some individuals, especially as the drug’s effects wear off.
A 2014 study by Kafka et al. investigated the anxiogenic effects of mixed amphetamine salts on developing male rats. Beginning at postnatal day 24, the rats were administered a mixture of chocolate drink and D,L-amphetamine at a dose of 1.6 mg/kg over 36 days.
The researchers repeatedly assessed anxiety responses immediately after daily amphetamine administration in the open field test, the social interaction test, and elevated plus maze. While no group differences were seen in the first two tests, the researchers noted significant anxiogenic responses in the elevated plus-maze. This maze tests basic approach and avoidance behavior and is commonly used to measure anxiety-related behaviors in rodents.
Specifically, the amphetamine-administered group demonstrated increased avoidance of the open arm entries, seen as reduced entries into the open arms compared to controls. Rodents generally prefer to avoid open spaces in novel environments because it is associated with being unsafe. However, a relatively high amount of avoidance is indicative of anxious behavior.
These findings by Kafka and colleagues are in agreement with earlier studies that have demonstrated the anxiogenic effects of amphetamine. A similar study in mice conducted by Lapin found that acute amphetamine administration dose-dependently reduces the number of entries into the open arms and total time spent in the open arms within the elevated plus-maze.
Adderall and Relapse Behavior in Adolescent Rats
Adderall addiction is associated with a high rate of relapse (reinstatement behavior), even after long periods of abstinence. The conditioned place preference paradigm is a widely used animal model to assess relapse behavior. In this model, the subject is trained to acquire a drug-induced conditioned place preference, and then subsequently goes through an extinction of the preference via the elimination of the compartment-drug association. Following extinction, reinstatement of the conditioned place preference (relapse behavior) can be assessed through the presentation of the drug at different time periods.
A 2008 study by Cruz et al. evaluated D,L-amphetamine-induced place conditioning in adolescent rats (postnatal day 42) and its subsequent reinstatement after extinction. To investigate reinstatement behavior, the researchers administered a priming injection of D,L-amphetamine (2.5mg/kg) one day, 30 days, and 60 days after place conditioning extinction. Reinstatement behavior was seen as an increase in time spent in the amphetamine-paired compartment compared to the neutral (saline) compartment.
They found that the priming injections of amphetamine were able to reinstate the conditioned place preference one day and 30 days, but not 60 days after extinction. The researchers concluded that amphetamine-induced place conditioning persists through early adulthood, corroborating other findings that adolescent psychostimulant exposure creates long-lasting neuroadaptations that render the subject more prone to relapse in the future.
To investigate neuromolecular alterations that may account for relapse behavior, Cruz and colleagues immediately dissected the nucleus accumbens after reinstatement. The nucleus accumbens is a key region of the mesolimbic dopamine system that plays a role in motivating responses to rewarding stimuli. The researchers observed decreased levels of Glutamate Receptor 1 (GluR1) on reinstatement days 1 and 30, suggesting that amphetamine-induced neuroplasticity on glutamatergic receptors in dopaminergic reward pathways can mediate relapse to drug-seeking behavior.
Pharmaceutical Interventions Counteracting Adderall’s Effects
Adderall’s Behavioral Effects Subdued by Diazepam
Benzodiazepines are commonly prescribed GABA agonists for anxiety, epilepsy, and sleep disorders. This class of drugs is known to reduce dopamine release in the nucleus accumbens, a crucial brain region in the mesolimbic dopamine reward pathway. With that knowledge, Guaita and colleagues aimed to see if diazepam would block the rewarding effects produced by D,L-amphetamine.
The researchers tested whether diazepam could block hyperlocomotion effects and ultrasonic vocalizations (USVs), particularly in the 50Khz range. USVs in this range are commonly produced by rats from rewarding situations such as food, sex, play, and the administration of dopaminergic psychostimulants.
First, the researchers found that D,L-amphetamine alone (dosed at 3mg/kg) caused significant increases in the number of USV calls, stereotypies (repetitive behaviors such as sniffing and rearing), and locomotor activity compared to the administration of the control, saline. Then, the researchers co-administered D,L-amphetamine with 2mg/kg of diazepam. They found that diazepam blocked the increase in the number and total duration of the 50Khz calls in addition to the stereotypy induced by D,L-amphetamine. However, diazepam did not block the increase in horizontal locomotion produced by D,L-amphetamine.
Guatita and colleagues also tested whether co-administration of amphetamine with haloperidol would blunt amphetamine’s effects. Haloperidol is a typical antipsychotic that antagonizes dopamine D2 receptors. Similar to the effects of diazepam, they found that haloperidol at a dose of 0.2mg/kg blocked 50-KHz USVs and stereotypies. Furthermore, haloperidol was able to subdue the increased locomotor activity caused by D,L-amphetamine, something which was not observed in the diazepam-treated experimental mice.
The researchers conclude that diazepam and haloperidol can effectively block the rewarding effects of amphetamine through its ability to reduce dopamine release in the mesolimbic dopamine reward circuitry. This is in agreement with previous studies demonstrating benzodiazepine’s ability to block reward-motivating tasks, such as in conditioned place preference and self-administration paradigms. The authors also note that the GABAA benzodiazepine receptor may, therefore, be an effective target to treat psychostimulant abuse. However, benzodiazepines carry their own potential for addiction.
Quetiapine Normalizes Long-Term Anxiolytic Effects Caused By a High-Dose Regimen of Adderall
Quetiapine is an atypical antipsychotic drug used to treat schizophrenia, bipolar disorder, and depression in humans. Similar to haloperidol, quetiapine is a dopamine D2 receptor antagonist, but also antagonizes serotonin and adrenergic receptors.
A 2005 study by He et al. investigated whether chronic administration of quetiapine would alleviate anxiety-like behavioral consequences of a neurotoxic amphetamine regimen given to rats.
First, the researchers characterized the behavioral profile of a high dose regimen of D,L-amphetamine given without quetiapine. They found that D,L-amphetamine (20mg/kg/day for 5 days) produced acute hyperthermia and reduced levels of anxious behavior when tested two weeks later. This was indicated by more time spent in: the unfamiliar part of the open field test as well as an increase in time spent in the light box in the light/dark box test, and in the open arms of the elevated plus-maze test. With this in mind, they wanted to see if quetiapine could alleviate these D,L-amphetamine-induced behavioral changes, possibly by bestowing neuroprotective effects on dopaminergic and serotonergic systems.
Compared to rats given D,L-amphetamine alone, the researchers found that quetiapine reduced the short-term increase in body temperature produced by D,L-amphetamine. Additionally, quetiapine normalized the long-term anxiolytic behavioral changes caused by D,L-amphetamine, lowering them down to control levels.
Overall, these findings differ from previous studies that have shown amphetamine produces anxiogenic responses, albeit in doses ranging from 0.5-5mg/kg. The authors conclude quetiapine is an effective therapeutic approach to regulate emotional changes caused by a neurotoxic regimen of amphetamine.
Adderall enhances monoamine neurotransmission by promoting reverse transport and blocking the reuptake of monoamine transporters. These studies demonstrate its wide-ranging effects on mood and behavior, namely, its anxiogenic properties, its effects on ADHD-like behavior, and its high potential for drug-induced relapse. In addition, benzodiazepines and antipsychotics have been found to attenuate Adderall’s rewarding and anxiety-like behavioral effects, respectively.
More rodent studies utilizing mixed amphetamine salts are needed to further delineate Adderall’s effects on brain physiology and behavior, especially when taken chronically.
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