Effects of L-theanine on Motor Cortex Excitability in Healthy Subjects: A Paired-Pulse TMS Study

Background and Significance:

Major depressive disorder (MDD) is a serious mental illness and the leading cause of disability worldwide. Although many antidepressants acting on synaptic monoamine levels have been used as the first-line drug treatment for MDD, around one third of MDD are pharmacologically resistant. Side effects of these medications impose additional hardship on adherence and further affect treatment outcome. New pharmacotherapeutic agents with complementary neurobiological mechanism and better side effect profile are of great needs. In addition to the monoamine system, the glutamatergic system plays a crucial role in MDD.

L-theanine (N5-ethyl-L-glutamine) is the primary psychoactive component uniquely in green tea. Epidemiological studies support that green tea consumption is an independent factor associated with lower prevalence of depression. Preclinical studies have demonstrated anti- depressant effect of L-theanine in rodents and provided evidences for its pharmacological properties of N-methyl-D-aspartate (NMDA) and gamma-aminobutyric acid (GABA) agonism. Yet these effects have not been proven in humans. Only one open-label clinical trial has studied and supported antidepressant effects of L-theanine in MDD patients. We propose using pair-pulse transcranial magnetic stimulation (ppTMS) to probe how L-theanine may manipulate the glutamatergic and GABA systems in the frontal region by changing cortical excitability first in healthy subjects. ppTMS is a well-established technique to investigate frontal motor cortical excitability mediated by the inter-neuron NMDA and GABA receptors. Specific changes of ppTMS measures, including impaired short-term and long-term intracortical inhibition (SICI, mediated by GABA-A receptor; LICI, mediated by GABA-B receptor) and intracortical facilitation (ICF, mediated by NMDA receptor), have been demonstrated in MDD. Using this technique, we plan to investigate the neurobiological effects of L-theanine in healthy subjects first.

Granted that the first phase pilot trial provides neurophysiological evidence of L-theanine on motor cortex excitability in human subjects, next phases of studies on L-theanine in MDD patients cortical excitability could be justified. This will lay foundation for further exploration of L-theanine's potential as an augmenting agent for MDD in a placebo- controlled design.

Aims and Hypothesis:

Given the potential NMDA and GABA agonistic effects of L-theanine, we hypothesize that it increases intracortical inhibition and facilitation through enhancement of NMDA- and GABA-receptor mediated neurotransmission, in healthy subjects (N=10 to complete study).

Study Procedures:

Double-blinded, Randomized-order, Cross-over placebo-controlled to evaluate acute effect of single-dose L-theanine on motor cortex excitability by ppTMS in 10 healthy subjects.

Dose of L-theanine or placebo is 400mg. At baseline, subjects will be randomized to L- theanine or placebo group, then receive ppTMS protocol before drug administration. The ppTMS protocol is repeated after 30min of administration. Then subjects will return to clinic after 1 week free of any medications and repeat the above protocol with the second drug condition. Visual analog scale will be used to evaluate psychosomatic symptoms and wellbeing of the subjects pre- and post-each drug administration.

Data Analytic Plan:

Wilcoxon test will be used to compare the baseline-to-post-drug means of SICI, LICI and ICF measures. Prespecified covariates include age, sex, handedness, level of fatigue will be analyzed in linear regression model. The time-condition relationship with continuous dependent variables of ICI and ICF values will be evaluated by Mixed Effect Model. Two- sided P value < 0.05 is considered statistically significant.

Inclusion criteria

• Adult, aged between 18 and 65 years old;
• Able to read/speak English and give informed consent
• No current or history of Axis I psychiatric disorders by DSM-5.
• Free of psychotropic medication use

Exclusion criteria

• History of significant acute or chronic neurological or medical disorder or condition that increases risk for seizure with TMS;
• History of alcohol use disorder, nicotine dependence, adjustment disorder;
• History of allergic reactions to L-theanine or green tea;
• Pregnancy;
• Unable/unwilling to abstain from nutraceutical supplements and psychotropic agents during participation in the study
• Unable/ unwillingness to refrain from recreational substance use (e.g. alcohol or marijuana) during participation in the study;
• Meet criteria for exclusion from TMS or MRI procedures, including intracranial metal implants or nonremovable ferromagnetic items in the head/neck.