Extracellular Protons: a Novel Modulator of Tyrosine Hydroxylase Activity in Catecholaminergic Cells

Tyrosine hydroxylase (TH) plays a crucial role in the catecholamine biosynthetic pathway in the brain, which is vital to the production of key neurotransmitters in the brain. TH is the rate-limiting enzyme in the production of catecholamines, which includes neurotransmitters such as dopamine, norepinephrine, and epinephrine. This means that the activity of TH largely determines the rate at which these neurotransmitters are produced. In the brain, dopamine is particularly significant, and TH is responsible for the first step in its synthesis. Dopamine plays an important role in various brain functions, including reward, motivation, memory, attention, and regulation of body movements. Any changes in dopamine levels may cause neurological and psychiatric disorders. For instance, reduced dopamine production, due to impaired TH activity, is a hallmark of Parkinson’s disease, leading to motor symptoms like tremors and rigidity. On the other hand, dysregulation of dopamine is also associated with psychiatric conditions such as schizophrenia and depression. Moreover, TH activity in the brain is also involved in the body’s response to stress. Norepinephrine, which is synthesized through the action of TH, plays a key role in the brain’s stress response, influencing attention, arousal, and mood. Because of its importance, the activity of TH is tightly regulated through various mechanisms, including feedback inhibition by catecholamines, phosphorylation, and changes in gene expression. This regulation is crucial for maintaining appropriate levels of catecholamines in the brain. In a new study published in the peer-reviewed Journal ACS Chemical Neuroscience led by professor Kandatege Wimalasena and conducted by Viet Le, Maya Eldani, and MyLinh Truong from the Department of Chemistry and Biochemistry at Wichita State University in Kansas-USA conducted a series of detailed experiments to understand the effect of extracellular proton concentration ([H⁺] _o) on TH activity in catecholaminergic cells. Traditionally, TH activity was believed to be governed predominantly by intracellular mechanisms, particularly the phosphorylation/ dephosphorylation of specific serine residues in response to intracellular calcium levels. The authors, however, introduced a novel extracellular factor: [H⁺] _o as a significant regulator of TH activity, independent of intracellular calcium.

The researchers employed an innovative experimental approach, focusing on MN9D and PC12 catecholaminergic cells. These cells were incubated in media with varying pH levels to modulate the extracellular proton concentration. To assess the activation of TH, the researchers measured the levels of DOPA, a direct product of TH activity. They found that lower pH (higher [H⁺] _o  ) significantly increased DOPA levels in a concentration-dependent manner. This increase was observed to be independent of extracellular sodium, potassium, and calcium ions but dependent on extracellular chloride ions. The authors examined the role of specific ion exchangers in this process using inhibitors like DIDS (a chloride ion transporter inhibitor) and amiloride (a non-specific sodium/hydrogen exchanger inhibitor). These inhibitors helped to dissect the role of specific ion exchangers in [H⁺] _o  -mediated TH activation.  They also investigated the effects of [H⁺] _o  on intracellular pH  [H⁺] _i  and calcium levels ([Ca^2+]_i) using pH and Ca^2+-sensitive fluorescent dyes. They observed that increasing [H⁺]_o caused a corresponding increase in [H+]_i, which was dependent on extracellular chloride. Notably, [H⁺]_o did not significantly affect [Ca²⁺]_i levels. The authors highlighted the unexpected role of chloride ions in this novel regulatory mechanism. The inhibition of [H⁺] _o  -mediated TH activation by DIDS, a chloride ion transporter inhibitor, suggests a critical role for chloride ions, possibly in maintaining the electrochemical gradient essential for proton influx.

The team demonstrated that increased extracellular proton concentration leads to enhanced phosphorylation of specific serine residues on TH, which is crucial for its activation. This process appeared to be independent of changes in [Ca²⁺]_i and involved changes in the phosphorylation state of TH rather than changes in TH expression levels. These findings indicate a novel mechanism of TH regulation, where extracellular protons act as a regulatory signal. This mechanism is distinct from the traditionally understood intracellular mechanisms, like intracellular calcium-mediated phosphorylation.

In conclusion, the implications of professor Kandatege Wimalasena and colleagues’ study are profound. Firstly, it reshapes our understanding of TH regulation, introducing extracellular proton concentration as a key factor. This finding is particularly significant in understanding how neuronal activity and synaptic release, which can alter extracellular pH, might impact catecholamine synthesis directly. Secondly, the research potentially opens new avenues for therapeutic interventions in conditions where catecholamine regulation is disrupted, such as Parkinson’s disease. By targeting extracellular pH regulation, it might be possible to modulate TH activity and, consequently, catecholamine levels in specific neuronal populations.