Many interventions addressing developmental disorders in the medical sciences, psychology, and pediatric developmental programs use an interdisciplinary approach. The intervention used in this study with the four specific groups of participants differs from the traditional forms of medical support (medications, surgical procedures, orthotics, etc.), PT (physiotherapy), OT (occupational therapy), SP (speech therapy), cognitive-behavioral therapy, neurofeedback [1] massage therapy, hydrotherapy, dietary advice, alternative treatments such as Bobath Neuro-Developmental Techniques [2] Doman-Delacato [3] Vojta [4] , Respiratory Therapy, as well as many others.

Masgutova Neurosensorimotor Reflex Method (MNRI) addresses the neurosensorimotor aspect of early sensory-motor patterns and reflexes to support sensory-motor integration and neurodevelopment of children and adults with neurodeficits and learning challenges: CP, TBI [5] [6] [7] [8] , ASD [9] [10] Down syndrome [11] [12] [13] [14] [15] and other [16] [17]. MNRI therapy meets the ever-increasing demands for neurorehabilitation of individuals with impaired sensorimotor functions due to damage or dysfunctions in central nervous systems. Previous studies of the MNRI Method and its different sub-programs demonstrated a positive effect on immune markers [13] [18] [19] , neurophysiological functions [7] [8] and different developmental aspects including the regulation of behavior and emotions, language, and communication [9] [17] [20] [21].

This paper presents a study of the effectiveness of MNRI therapy using neurotransmitter analysis. The aim was to investigate whether the neurotransmitters markers improve as a result of MNRI Reflex Integration treatment modality for individuals with global developmental disorders, cerebral palsy, traumatic brain injury (TBI), acute brain injury (ABI), seizures, ADD/ADHD, and anxiety disorders.

The MNRI method was developed in 1989 in Russia and further elaborated in Eastern Europe to treat individuals with certain types of sensorimotor or reflex development deficits, behavior disorders, disorders of speech or language development, and learning disabilities. It was introduced to the USA in 1996 and has gradually become adopted in many other countries. Clinical observations showed that MNRI facilitates the neurodevelopment in individuals having various neurological deficits and enables them to reroute and improve their early movements, reflex patterns, coordination systems, and skills to enable more optimal functioning, development, and learning [5] [6] [7] [8] [9] [11] [13]. The MNRI therapy program is based on the theory that impaired reflex circuits can be reconstructed and re-integrated, which involves awakening the genetic sensorimotor memory in individuals even with severe diagnosis, for example, with CP and brain damage [5] [6]. MNRI is an evidence and research-based system of a therapeutic program oriented to analyze the effect of the MNRI reflex integration techniques for neurosensorimotor-cognitive development of children and adults with neurodeficits and learning problems [22]. The studies were conducted by Dr. S. Masgutova and her team and scientific colleagues from different countries between 1989 and 2016 [22].

MNRI is based on exercises and techniques called “repatterning”, meaning re-education, recoding, rerouting of and paving the reflex nerve pathways specific for dynamic and postural reflex schemes (e.g., Babinski, Automatic Gait, Bauer Crawling, Hands Grasp, and others) (Reflexes: Portal to Neurodevelopment and Learning 2015). The stimulation of reflex pathways is aimed at strengthening and stabilizing the traces of genetic sensory-motor memory and activation of innate defensive mechanisms of the body’s brain “alarm” system (HPA-stress axis or “hypothalamus- > pituitary gland- > adrenals” cycle activation) in times of stress or danger [23]. MNRI exercises stimulate innate neuro-regulation mechanisms and resilience in the stress and immune systems [8] [9] [24]. Repatterning activates the extra pyramidal nerve system (peripheral nerves, spinal cord, brain stem, and diencephalon) responsible for automatic mechanisms and processes, the extension of links between neurons, the growth of neural nets, myelination, and the creation of new nerve routing, as described by Sechenov [25] [26] , Pavlov [27] , Anokhin [28] , Haines [29] , Virella [30].

There are various ways to assess neurotransmitters in the body. The three most common ways to achieve this include:

1) The commonly accepted method of testing the neurotransmitters is cerebral spinal fluid (CSF). While this approach does accurately reflect what is happening with the central nervous system transmitter levels, there are considerable drawbacks to utilizing this method in an educational facility for and non-medical purposes. Collection of the spinal fluid can also create stress in the participant, therefore increasing levels of key neurotransmitters such as norepinephrine and epinephrine. Another drawback to CSF usage is that there is yet to be established optimal ranges for neurotransmitters, therefore it is difficult to have consistent and relevant interpretation.

2) Another option for testing is by venipuncture. This method also increases stress levels and therefore has the potential of altering the true neurotransmitter levels. Blood samples would require special handling to maintain the fluid’s integrity.

3) Given the invasiveness and instability issues of methods (1) and (2), urine testing provides the most optimal alternative.

For nearly 60 years, studies have utilized urinary measurements of neurotransmitters and neurotransmitter metabolites (Marc, et al. 2010). Since the 1950s, increased urinary excretion of dopamine (DA), norepinephrine (NE), and epinephrine (E), the three naturally occurring catecholamines, has been used to diagnose pheochromocytoma. Early days concerns about the limitations of urinalysis for neurotransmitters, were alleviated by development of fluorometric methods. High-Performance Liquid Chromatography (HPLC) methodology has expanded parameters of even greater specificity and sensitivity, allowing an even wider span of clinical application [31] [32] [33]. Lynn-Bullock examined the effects of oral ingestion of the serotonin substrate (5-hydroxytryptophan (5-HTP)) on specific brain regions in rats [34]. Serotonin is a monoamine neurotransmitter and is implicated in many physiological and behavioral functions including: affect, aggression, appetite, cognition, emesis, endocrine function, gastrointestinal function, motor function, neurotropism, perception, sensory function, sex, sleep, and vascular function [34] [35]. Serotonin levels were measured using brain tissue immunoreactivity and urinalysis and observed maximum serotonin immunoreactivity in the serotonergic dorsal raphe nucleus within 2 h of 5-HTP administration. Urinary analysis of serotonin, 5-HTP, and 5-hydroxyindolacetic acid (5-HIAA), a serotonin metabolite, had parallel changes in immunoreactivity to the dorsal raphe nucleus, demonstrating a positive correlation between CNS serotonergic activity and urinary serotonin levels [35] [36].

Studies have further determined that those struggling with depression, ADD, and ADHD have a direct correlation to neurotransmitters in the CNS and the urine. There are several reasons preventing the use of urinary neurotransmitter levels as a sole diagnostic tool. However, the utility of urinary neurotransmitter levels as indicators and biomarkers to gauge the realm of impact and treatment is unquestionable [37]. Clinicians are already utilizing urinary histamine markers as an accurate assessment tool and treatment directive. Studies have also demonstrated a direct correlation between neurotransmitter levels and acute PTSD in children [38].

This manuscript focused on the testing of the following neurotransmitters: epinephrine (1), norepinephrine (2), dopamine (3), DOPAC (4), serotonin (5), 5-HIAA (6), glycine (7), taurine (8), GABA (9), glutamate (10), PEA (11), and histamine (12).

Some of the studied neurotransmitters are related biochemically: 5HIAA is a metabolite of serotonin, and DOPAC is a metabolite of dopamine. The neurotransmitters can be divided into two categories by: inhibitory vs. excitatory. This classification is complicated, since transmission of neurotransmitters is a complex system and any neurotransmitter can have a secondary function. For instance, GABA synapses are typically assumed to be inhibitory, however inhibitory and excitatory GABA connections coexist in the cerebellar interneuron network. Psychoneuroimmunology studies have shown that the central nervous system has complex interactions with the endocrine and the immune systems. Although the mechanisms of these interactions are not fully understood, it is known that stress can induce modulation of the immune system, acting via the hypothalamic-pituitary-adrenal circuit and the sympathetic-adrenal medullary axes [23] [29] [30] [39].

Immune and central nervous systems are integrative and interdependently maintaining. The central nervous system modulates the immune system by hardwiring sympathetic and parasympathetic nerves (autonomic nervous system) to lymphoid organs. Neuroendocrine hormones (such as corticotrophin-releasing hormone or substance P) regulate the cytokine balance. The immune system, in turn, affects brain activity including inhibition-excitation processes, as well as sleep, and body temperature. Working in this highly reciprocal manner the immune-nervous system creates a unique functional and anatomical link for the organism to act at multiple levels [29] [39]. Stress can disrupt interaction between the nervous and immune system [39].

There are three major forms of neurotransmitters functioning in a human organism, all of which communicate via signals targeting specific cells. These are: acetylcholine neurotransmitter, biogenic amines, and amino acids. Below we present a general overview of the analyzed transmitters.

Gamma Amino Butyric Acid (GABA) is the major inhibiting or calming transmitter in a human organism. The role of GABA is to inhibit or reduce the activity of the neurons or nerve cells, and the calming effect is achieved by restricting neuroelectric activity [40]. The body relies on GABA to reduce anxiety. The activity of GABA is increased during sleep, making GABA responsible for setting the circadian rhythms in humans. Other functions are impacted by GABA, for example, female hormonal cycles are affected by circadian rhythm. GABA is prominent in the hippocampus, this part of the brain is responsible for memory formation, by allowing the brain to reconfigure neural connections [40] [41]. Excess of GABA may lead to depression or apathy; low levels of GABA are known to affect sleep. There is also a potential influence of GABA on immunity and physiology wellness [42].

Glutamate is an excitatory neurotransmitter that enhances signal transduction; it is tied to learning and memory; it is also one of the most common neurotransmitters: there is more glutamate in the brain than any other amino acid. Tissue extracted from the brain contains 5 - 15 mmol glutamate per kg [43]. Glutamate is not only involved in cognitive functions, but also learning function. Being an amino acid, it has a role in metabolic function and ammonia detoxification. Traumatic brain injury or stroke may create physical damage which in turn causes glutamate to bring hyper excitation and thereby killing neurons, thus resulting in brain damage [44].

Norepinephrine assists the body to go into “high alert” and is responsible for increasing heart rate and blood pressure. Norepinephrine can activate not only pre but post synaptic adrenergic receptors. Norepinephrine is released in the brain as the locus coeruleus is stimulated and action potential occurs. It is also important for the long-term memory by influencing surges in specific neural circuits. When norepinephrine binds to a receptor and stimulates a nerve to respond, it works as a neurotransmitter. When it is originated in the adrenal medulla as a product of dopamine, it works as a hormone (Epinephrine and Norepinephrine, https://www.boundless.com/blog/biology/).

Epinephrine is another flight/fight hormone released when the body is under extreme stress. It is closely related to norepinephrine since it is synthesized from norepinephrine in the adrenal medulla. Norepinephrine and epinephrine act on beta and alpha receptors, the difference is that norepinephrine is primarily focused on alpha receptors, whereas epinephrine stimulates all three beta receptors as well as alpha receptor.

Dopamine is the precursor molecule to both norepinephrine and epinephrine in the catecholamine synthesis pathway. This important neurotransmitter is produced several areas of the brain. The midbrain is host to the substantia nigra, which is a part of the basal ganglia and plays an important role in the reward system, voluntary motor movements, cognition and emotions. High levels of neuromelanin in dopaminergic neurons make substantia nigra appear darker than surrounding areas [45]. One function of dopamine is movement; the caudate and putamen create the striatum, which is a bundle of dopaminergic neurons that form the nigrostriatal pathway considered a facilitator of movement. The second function is that of the reward system, which originates in the ventral tegmental area of the brain. Wolfram Schultz’s 1990 experiment demonstrated that dopamine release was associated more with unpredictability rather than with mere experience or a reward system. There are two main metabolites of dopamine: Norepinephrine and DOPAC, which is correlated with oxidative stress.

Serotonin (produced in the enteric nervous system of the gastrointestinal tract and in the central nervous system) has many important roles in overall health and wellbeing. It is a chemical product of nerve cells and serves in the modulation of glutamate excitation through voltage potential regulation. Serotonin is a metabolite of 5-HTP; up to 90% of serotonin is produced in the gastrointestinal tract, although it is also found in platelets and throughout the central nervous system. Serotonin is involved with smooth muscle contraction, regulating cyclic body processes and nerval impulse transmission. Most agree serotonin is the primary neurotransmitter for mood regulation [46]. Serotonin must originate in the brain if it is to be utilized by the brain. It is widespread in its roles and influences mood, digestion, ambition, social behavior, memory and sexual function. Serotonergic hypo function has been demonstrated to be a neurochemical trait associated with or precursor to impulsivity and aggression [46]. Serotonin also acts as a precursor to melatonin, which plays a major role in the body’s circadian rhythm and sleep cycles. Both dopamine and serotonin have a close relationship in mood, anxiety, and aggression. The inhibitory and excitatory relationship between these two neurotransmitters has been the subject of numerous studies [46] [47] [48]. The main metabolite of serotonin is 5-HIAA. In our study, we analyzed 5-HIAA to obtain insight into the production of serotonin and the potential absorption concerns due to the tryptophan metabolites.

Taurine is a trophic factor and assists in the survival of synaptic neurons. Taurine is a neuromodulator and is protective against glutamate-induced excitotoxicity through limiting increase of intracellular calcium levels. This is achieved by shifting the ratio of BCL-2 and BAD ratio in favor of cell survival and reducing endoplasmic reticulum stress. Taurine plays an integral part in regulation of the amount of sensory information received by the cortex. Taurine is partly responsible for helping the brain to turn off at night by activating GABA, it is stopping the thalamus (processing input from the senses) from waking up a sleeping person [49] [50]. Taurine is also involved in maintaining metabolic homeostasis in the body by reducing blood glucose levels and helps restore insulin sensitivity. Studies have demonstrated that taurine can reverse tinnitus, since many issues with hearing are a result of nerve cell communication with the brain as opposed to mechanical issues. Taurine assists in calcium restoration and control of auditory cells. Taurine is also involved in cortisol elimination, therefore mitigating stress responses within the body [49] [51] [52].

Histamine has many physiological roles within the body, one of which is the ability to increase excitability of the central nervous system. Studies have demonstrated that histamine can be a regulator of “whole brain” activity. Studies have also confirmed the potential of histamine to assist in mediation of alertness and electrographic arousal [53] [54] [55]. The hypothalamus’s ventral preoptic center assists in turning off histaminergic tuberomammillary cells which connects to the sleep/wake cycle resulting in diurnal changes in brain function. Many functions of the hypothalamus are regulated by histamine. Vasopressin is physiologically regulated by histaminergic neurons [54] [55]. Additional studies demonstrate the potential of histamine for regulating levels of ACTH, prolactin, and oxytocin [55].

Glycine is another inflammation regulator; it plays an important role in overall health and wellness of the body. Glycine can be produced by the liver in limited amounts and is used for detoxification. Glycine helps with synthesis of bile acid, amino acids, and assists in the building of DNA, RNA and protein. By being involved in the manufacturing of immune-related hormones, it also plays a role in immunological function. Glycine is found in the spinal cord, brain stem, retina, and throughout the central nervous system. It acts on inflammatory cells such as macrophages to suppress activation of transcription factors and the formation of free radicals and inflammatory cytokines. Glycine is not just an anti-inflammatory agent, but also an immunomodulator and cytoprotective agent [56]. Excessive amounts of glycine can lead to decreased energy, anxiousness, sleep difficulties, as well as immune dysregulation and digestive stress.

PEA (phenylethylamine) is an excitatory neurotransmitter that assists in modulation of action potentials to enhance glutamate activity and neurotransmitter firing. It has been demonstrated that amongst the ADHD population there were consistently lower levels of PEA and PEA metabolites [57]. PEA has a tremendous impact on overall body alertness and physiological performance. Emotions are regulated in limbic region and having a very high concentration of PEA makes an impact on motivation, emotions and socialization. Because it is an endogenous stimulant, PEA enhances dopamine, norepinephrine and serotonin. PEA causes fast impulse-mediated release of catecholamines (dopamine and epinephrine) and serotonin in the brain, resulting in rapid improvement of cognitive performance, attention, awareness, pleasure, libido, and overall sense of wellbeing [58]. Furthermore, increase in PEA improves alertness and higher cognitive functioning [48]. PEA can increase catecholamine levels and block re-uptake of dopamine, norepinephrine, and serotonin. This can bring restoration of energy as well as mood elevation. However, too much PEA can lead to anxiety. Studies demonstrate that those with ADHD will show either low or high levels of PEA. High levels can also be associated with anxiety, insomnia and mind racing [57] [59].

The baseline data on analysis of neurotransmitters markers for individuals with global developmental disorders (Group 1), cerebral palsy, TBI, and seizures (Group 2), ADD and ADHD (Group 3), and anxiety disorders (Group 4) are presented in Table 2. The effect size was assessed using Cohen’s ɗ (see Methods) to determine the change in neurotransmitter levels between pre- and post-treatment intervention.

Our study demonstrated the impact on the neurological system from the MNRI treatment. The tested neurotransmitters are chemical messengers in our brain and body, delivering signals from one synapse to another. To ensure proper transmission from the presynaptic cell to the receptors of the postsynaptic cell the neurotransmitters must have appropriate genetic and chemical structure [29] [64]. Neurotransmitters participating in inhibitory or excitatory synapses determine functional properties of the underlying circuit. Depending on the specific pair of neurotransmitters and corresponding receptors, qualitative and quantitative features of the synapse vary due to the distinct kinetics and signaling cascades imprinted in the identity of the synaptic partners [64] [65]. This mechanism is determined by function of sensory neurons and, on other side, defines

characteristics of the processing of the input on level of neurotransmitter specification [29] [66] [67].

We have tested the following neurotransmitters: epinephrine (1), norepinephrine (2), dopamine (3), DOPAC (4), serotonin (5), 5-HIAA (6), glycine (7), taurine (8), GABA (9), glutamate (10), PEA (11), and histamine (12). Below the interpretation of neurotransmitter levels in pre- and post-intervention MRNI therapy for all study group (n = 80) individually compared to the Healthy Control group (n = 117) is presented.

The present findings suggest that the MNRI treatment positively impacts central nervous, immune, and endocrine systems by regulation of the stress hormones and neurotransmitters regulation. The results indicate that through treatment using reflex patterns there is a reduction in the excitability within the nervous system, and reduction in the inflammatory response. All four study groups demonstrated positive changes that were generally small scale. The anxiety disorders group showed medium effect in the histamine response, and a large effect on the neurotransmitter taurine. The ADD/ADHD group had remarkable medium-to-large effects in most neurotransmitters. Neurotransmitter regulation can influence mood, behavior, focus, and cognitive functions. By regulation of the neurotransmitters and increasing the neuromodulation effect, the pathway for neurodevelopment, increase of stress resilience, neuroplasticity and new learning is facilitated. It is pertinent to note that modulation is achieved without the use of pharmaceuticals and their attendant risk of side effects.

The study was conducted during the MNRI Family Conferences across the United States.

This study was approved by New England IRB (#: 20160464, Legacy IRB #: 15-466, Action Date: 02/11/2016; A WIRB Copernicus Group Company). All specialists leading the evaluations were certified by the NIH (National Institute of Health, Office of Extra Mutual Research) “Protecting Human Research Participants” in 2012-2017. All participants were assigned codes to protect anonymity. Receipt of informed consent was received from all participants’ parent or legal guardian. MNRI Therapy Program was conducted and treatment administered by designated Specialists or MNRI Core Specialists who have successfully completed the requirements for Continuing Professional Education in MNRI and clinical hours (www.MasgutovaMethod.com). Double blind analysis of research results and preparation of this report is done by independent experts.

The authors declare no conflicts of interest regarding the publication of this paper. The scientists performing the qualitative and quantitate analysis of these research results were not privy to details regarding the treatment or suspected outcomes. They also had no vested interest in MNRI.

Bell, C., Whitney, J., Deiss, T., Tatarinova, Т., Franckle, L., Beaven, S. and Davis, J. (2019) The Effect of the MNRI Method on Neurotransmitter Biomarkers of Individuals with Neurodevelopmental Disorders. Neuroscience & Medicine, 10, 292-321. https://doi.org/10.4236/nm.2019.103022