**8. The importance of dopamine and other neurotransmitters**

As already noted, some experimental animal and human studies concluded that behavioral changes may be explained by increased dopamine levels in the parasitized brain, and that these disorders could largely be resolved by administration of a dopaminergic receptor antagonist (e.g., haloperidol) or dopamine reuptake inhibitor (e.g., GBR‐12909) [96, 114, 115]. It is therefore possible that dopamine represents the link between toxoplasmosis and schizophrenia [97]. This neurotransmitter is synthesized in the cytosol of neurons from l‐tyrosine amino acid by the action of the tyrosine hydroxylase enzyme, which converts it to l‐3,4‐dihydroxyphenylalanine (l‐DOPA). l‐DOPA is in turn converted by the action of DOPA‐decarboxylase (DDC) to dopa‐ mine, a precursor of norepinephrine (noradrenalin) and epinephrine (adrenalin) in the synthe‐ sis pathway of these catecholamines. It is subsequently packaged in vesicles and transported through the axon to the synapse, where it is released by exocytosis in response to an electri‐ cal stimulus. Dopamine is one of the main neurotransmitters in the prefrontal cortex and the mesolimbic system (mainly formed by the nucleus accumbens, amygdala, and hippocampus), where the presence of *T. gondii* tissue cysts is especially frequent [73].

The definitive mechanism by which *T. gondii* induces changes in the dopaminergic pathway has not been fully elucidated. However, an increase in dopamine with no modification of cellular tyrosine hydroxylase was demonstrated *in vitro* after parasitization of a rat pheochro‐ mocytoma cell line (PC12) and *in vivo* after the parasitization of mouse brains. This dopamine synthesis is attributable to the additional activity of the aromatic amino acid hydroxylase, which is encoded by two *T. gondii* genes [116] and has homologous activities to those of mam‐ malian tyrosine hydroxylase, associated with the entry of cellular DDC enzymes into parasi‐ tophorous vacuoles (compartments formed by the parasite to invade the cell) and into tissue cysts (the protozoan encodes no enzyme with DDC activity) [114, 117]. Experiments in cell cultures have demonstrated that dopamine increases the replication of *T. gondii* tachyzoites [118]. This biochemical mechanism may play a role in the behavioral changes observed, which would result from the involvement of catecholaminergic neurons and consequent dopami‐ nergic hyperactivity [19].

Parasitization in the fetal period may also impair the development of mesolimbic dopaminer‐ gic or prefrontal cortex neurons (inappropriate migration, altered position, reduced synapses, etc.) leading to neurodevelopmental disorders. Disease symptoms would not be induced immediately by these early anomalies but would rather manifest after a latency period of one to three decades. This is because the proliferation, migration, differentiation, and maturation of glial progenitor cells continue throughout childhood [119] and the volume of gray matter increases to a peak in puberty before beginning to diminish [120].

However, the hypothesis that increased dopamine levels or dopaminergic hyperactivity is the underlying cause of schizophrenia does not account for the negative symptoms in these patients, which are more likely to result from dopaminergic hypoactivity. Therefore, neu‐ rotransmitters other than dopamine may play an important role in the development of this disease. Thus, it has been proposed that deficits in glutamatergic brain systems also partici‐ pate in the physiopathology of schizophrenia based on findings of higher kynurenic acid levels in patients with psychotic symptoms than in healthy controls [121]. Kynurenic acid is a metabolite of tryptophan with important biological effects on the nervous system, related to its antagonism for the glutamate receptor in the human brain (it is a glutamatergic NMDA receptor antagonist). Increased kynurenic acid levels due to blockade of NMDA receptors in glutamatergic neurons have been related to changes in dopamine level in different brain areas. These modifications include cortical dopaminergic hypoactivity and mesolimbic dopa‐ minergic hyperactivity, which would explain the negative and positive symptoms in patients. This relationship between glutamate pathway disorders and dopamine level changes may explain the presence of different symptoms in the psychosis [122].

expected that a high degree of degenerated neuron degeneration and cognitive impairment is

As already noted, some experimental animal and human studies concluded that behavioral changes may be explained by increased dopamine levels in the parasitized brain, and that these disorders could largely be resolved by administration of a dopaminergic receptor antagonist (e.g., haloperidol) or dopamine reuptake inhibitor (e.g., GBR‐12909) [96, 114, 115]. It is therefore possible that dopamine represents the link between toxoplasmosis and schizophrenia [97]. This neurotransmitter is synthesized in the cytosol of neurons from l‐tyrosine amino acid by the action of the tyrosine hydroxylase enzyme, which converts it to l‐3,4‐dihydroxyphenylalanine (l‐DOPA). l‐DOPA is in turn converted by the action of DOPA‐decarboxylase (DDC) to dopa‐ mine, a precursor of norepinephrine (noradrenalin) and epinephrine (adrenalin) in the synthe‐ sis pathway of these catecholamines. It is subsequently packaged in vesicles and transported through the axon to the synapse, where it is released by exocytosis in response to an electri‐ cal stimulus. Dopamine is one of the main neurotransmitters in the prefrontal cortex and the mesolimbic system (mainly formed by the nucleus accumbens, amygdala, and hippocampus),

The definitive mechanism by which *T. gondii* induces changes in the dopaminergic pathway has not been fully elucidated. However, an increase in dopamine with no modification of cellular tyrosine hydroxylase was demonstrated *in vitro* after parasitization of a rat pheochro‐ mocytoma cell line (PC12) and *in vivo* after the parasitization of mouse brains. This dopamine synthesis is attributable to the additional activity of the aromatic amino acid hydroxylase, which is encoded by two *T. gondii* genes [116] and has homologous activities to those of mam‐ malian tyrosine hydroxylase, associated with the entry of cellular DDC enzymes into parasi‐ tophorous vacuoles (compartments formed by the parasite to invade the cell) and into tissue cysts (the protozoan encodes no enzyme with DDC activity) [114, 117]. Experiments in cell cultures have demonstrated that dopamine increases the replication of *T. gondii* tachyzoites [118]. This biochemical mechanism may play a role in the behavioral changes observed, which would result from the involvement of catecholaminergic neurons and consequent dopami‐

Parasitization in the fetal period may also impair the development of mesolimbic dopaminer‐ gic or prefrontal cortex neurons (inappropriate migration, altered position, reduced synapses, etc.) leading to neurodevelopmental disorders. Disease symptoms would not be induced immediately by these early anomalies but would rather manifest after a latency period of one to three decades. This is because the proliferation, migration, differentiation, and maturation of glial progenitor cells continue throughout childhood [119] and the volume of gray matter

However, the hypothesis that increased dopamine levels or dopaminergic hyperactivity is the underlying cause of schizophrenia does not account for the negative symptoms in these

associated with the presence of *T. gondii* in the brain [111].

68 Schizophrenia Treatment - The New Facets

**8. The importance of dopamine and other neurotransmitters**

where the presence of *T. gondii* tissue cysts is especially frequent [73].

increases to a peak in puberty before beginning to diminish [120].

nergic hyperactivity [19].

Indoleamine‐2,3‐dioxygenase and therefore the metabolism of tryptophan, a precursor metab‐ olite in serotonin (and melatonin) synthesis, are induced by the proinflammatory cytokines released in response to *T. gondii* infection, especially IFNγ [123]. Tryptophan is an essential amino acid for the parasite, and decreased levels inhibit its growth and replication capac‐ ity [124]. However, induction of this metabolite in turn increases kynurenic acid levels and therefore alters dopamine levels through the glutamatergic receptor antagonist effect of this acid [114]. Tryptophan degradation also reduces serotonin levels, which has been related to a higher incidence of depression and suicide [125, 126], as also observed in patients with high anti‐*T. gondii* antibody levels [56].

Patients with schizophrenia also show anomalous levels of gamma‐aminobutyric acid (GABA), another important neurotransmitter [20], which is synthesized from glutamate by the action of glutamic acid decarboxylase (GAD) [127]. GABA activates GABAA receptors, which are ion channels, and GABAB receptors, which are G‐protein‐coupled receptors [128]. It is the main neurotransmitter with inhibiting effect in the CNS, regulating dopaminergic activ‐ ity and playing a key role in the reduction of neuronal excitability throughout the nervous system. Dopaminergic neurons in basal ganglia would be directly inhibited by GABAergic neurons, so that any GABAergic hypofunction would be accompanied by an increase in subcortical dopaminergic activity, as observed in schizophrenia.

More direct evidence of the involvement of this neurotransmitter in the etiology of schizo‐ phrenia derives from data on the reduction in neurons in the GABAergic system or in brain regions such as the hippocampus, temporal lobe, and prefrontal cortex of schizophrenic patients [129–131]. *Postmortem* molecular studies have demonstrated: a reduction in messen‐ ger RNA (mRNA) levels of isoform 67 of glutamic acid decarboxylase (GAD67) and of type 1 GABA transporter (GAT‐1) in the prefrontal cortex of schizophrenics [132, 133]; an increase in subunit α2 of the GABAA receptor in the initial segment of the axon of pyramidal neurons [134]; and a reduced expression of the receptor GABAB, which regulates GABA release as a possible compensatory mechanism for GABAergic dysfunction [135]. As noted above, these findings may be the consequence of alterations during neurodevelopment in the differentia‐ tion and migration of these neurons toward their definitive localizations in the brain. This would give rise to structural alterations and neurochemical dysregulation that would have a global effect on all of these neurotransmitters (dopamine, glutamate, serotonin, GABA) and would become manifest from adolescence onward, inducing the appearance of the disease. Once more, infection by *T. gondii* may play an important role in this process.

Outside the nervous system, GABAergic mechanisms have been observed in different tissues and peripheral organs, and GABA has also been found to exert a major role in the immune system, with important inter‐regulatory functions between this and the CNS [136]. It has been reported that *T. gondii* infection is followed by an increase in the motility and migratory capacity of infected dendritic cells, permitting propagation of the parasite to different tissues, including the brain [128]. Although dendritic cells are considered guardians of the immune system, they can also, paradoxically, mediate in the spread of the parasite. This mechanism is produced by the induction in these cells of the GAD enzyme and therefore of GABA pro‐ duction and secretion, which in turn activate GABA receptors expressed by these same cells, stimulating their motility [137]. In experimental mouse models, inhibition of the GABAergic pathway by blockade of GABAA receptors or inhibition of the GAD enzyme markedly reduced the hypermotility and spread of *T. gondii*‐infected dendritic cells and therefore of the parasite itself [137, 138]. Finally, it has also been reported that brain infection by *T. gondii* can interfere with the GABAergic system by inducing changes in the distribution of the GAD67 enzyme, although this event has been related more to possible neurological complications of toxoplasmic encephalitis, such as seizures [139], than to possible complications of latent toxoplasmosis, such as schizophrenia.

Accordingly, the inflammatory response of the host to parasitization, which aims to control parasite replication and alterations in differentiation and migration processes, can change levels of dopamine, tryptophan, kynurenic acid, serotonin, and GABA, leading to behavioral changes and giving rise to different psychotic symptoms.

In order to establish dopamine and other related neurotransmitters as a causal link between toxoplasmosis and schizophrenia development, it is necessary to confirm that this neurotrans‐ mitter is also involved in the disease genesis when there is infection by other pathogens [140], and this mechanism should also explain the possible contribution of *T. gondii* parasitization in other dopaminergic pathway diseases, e.g., Parkinson's disease [114].
