Elevated CYP2C19 expression is associated with depressive symptoms and hippocampal homeostasis impairment.

This corrects the article DOI: 10.1038/mp.2016.204.

Elevated CYP2C19 expression is associated with depressive symptoms and hippocampal homeostasis impairment.

The polymorphic CYP2C19 enzyme metabolizes psychoactive compounds and is expressed in the adult liver and fetal brain. Previously, we demonstrated that the absence of CYP2C19 is associated with lower levels of depressive symptoms in 1472 Swedes. Conversely, transgenic mice carrying the human CYP2C19 gene (2C19TG) have shown an anxious phenotype and decrease in hippocampal volume and adult neurogenesis. The aims of this study were to: (1) examine whether the 2C19TG findings could be translated to humans, (2) evaluate the usefulness of the 2C19TG strain as a tool for preclinical screening of new antidepressants and (3) provide an insight into the molecular underpinnings of the 2C19TG phenotype. In humans, we found that the absence of CYP2C19 was associated with a bilateral hippocampal volume increase in two independent healthy cohorts (N=386 and 1032) and a lower prevalence of major depressive disorder and depression severity in African-Americans (N=3848). Moreover, genetically determined high CYP2C19 enzymatic capacity was associated with higher suicidality in depressed suicide attempters (N=209). 2C19TG mice showed high stress sensitivity, impaired hippocampal Bdnf homeostasis in stress, and more despair-like behavior in the forced swim test (FST). After the treatment with citalopram and 5-HT1A receptor agonist 8OH-DPAT, the reduction in immobility time in the FST was more pronounced in 2C19TG mice compared with WTs. Conversely, in the 2C19TG hippocampus, metabolic turnover of serotonin was reduced, whereas ERK1/2 and GSK3ß phosphorylation was increased. Altogether, this study indicates that elevated CYP2C19 expression is associated with depressive symptoms, reduced hippocampal volume and impairment of hippocampal serotonin and BDNF homeostasis.

Ethanol self-administration and nicotine treatment increase brain levels of CYP2D in African green monkeys.

BACKGROUND AND PURPOSE: CYP2D6 metabolizes many centrally acting drugs, neurotoxins and endogenous neurochemicals, and differences in brain levels of CYP2D have been associated with brain function and drug response. Alcohol consumers and smokers have higher levels of CYP2D6 in brain, but not liver, suggesting ethanol and/or nicotine may induce human brain CYP2D6. We investigated the independent and combined effects of chronic ethanol self-administration and nicotine treatment on CYP2D expression in African green monkeys. EXPERIMENTAL APPROACH: Forty monkeys were randomized into control, ethanol-only, nicotine-only and ethanol + nicotine groups. Two groups voluntarily self-administered 10% ethanol in sucrose solution for 4 h·day(-1) , whereas two groups consumed sucrose solution on the same schedule. Two groups received daily s.c. injections of 0.5 mg·kg(-1) nicotine in saline bid, whereas two groups were injected with saline on the same schedule. KEY RESULTS: Both nicotine and ethanol dose-dependently increased CYP2D in brain; brain mRNA was unaffected, and neither drug altered hepatic CYP2D protein or mRNA. The combination of ethanol and nicotine increased brain CYP2D protein levels to a greater extent than either drug alone (1.2-2.2-fold, P < 0.05 among the eight brain regions assessed). Immunohistochemistry revealed the induction of brain CYP2D protein within specific cell types and regions in the treatment groups. CONCLUSIONS AND IMPLICATIONS: Ethanol and nicotine increase brain CYP2D protein levels in monkeys, in a region and treatment-specific manner, suggesting that CNS drug responses, neurodegeneration and personality may be affected among people who consume alcohol and/or nicotine.

Differential effects of nicotine treatment and ethanol self-administration on CYP2A6, CYP2B6 and nicotine pharmacokinetics in African green monkeys.

In primates, nicotine is metabolically inactivated in the liver by CYP2A6 and possibly CYP2B6. Changes in the levels of these two enzymes may affect nicotine pharmacokinetics and influence smoking behaviors. This study investigated the independent and combined effects of ethanol self-administration and nicotine treatment (0.5 mg/kg b.i.d. s.c.) on hepatic CYP2A6 and CYP2B6 levels (mRNA, protein, and enzymatic activity), in vitro nicotine metabolism, and in vivo nicotine pharmacokinetics in monkeys. CYP2A6 mRNA and protein levels and in vitro coumarin (selective CYP2A6 substrate) and nicotine metabolism were decreased by nicotine treatment but unaffected by ethanol. CYP2B6 protein levels and in vitro bupropion (selective CYP2B6 substrate) metabolism were increased by ethanol but unaffected by nicotine treatment; CYP2B6 mRNA levels were unaltered by either treatment. Combined ethanol and nicotine exposure decreased CYP2A6 mRNA and protein levels, as well as in vitro coumarin and nicotine metabolism, and increased CYP2B6 protein levels and in vitro bupropion metabolism, with no change in CYP2B6 mRNA levels. Chronic nicotine resulted in higher nicotine plasma levels achieved after nicotine administration, consistent with decreased CYP2A6. Ethanol alone, or combined with nicotine, resulted in lower nicotine plasma levels by a mechanism independent of the change in these enzymes. Thus, nicotine can decrease hepatic CYP2A6, reducing the metabolism of its substrates, including nicotine, whereas ethanol can increase hepatic CYP2B6, increasing the metabolism of CYP2B6 substrates. In vivo nicotine pharmacokinetics are differentially affected by ethanol and nicotine, but when both drugs are used in combination the effect more closely resembles ethanol alone.

Independent and combined effects of ethanol self-administration and nicotine treatment on hepatic CYP2E1 in African green monkeys.

Cytochrome P450 2E1 metabolizes ethanol and also bioactivates many toxins and procarcinogens. Elevated levels of hepatic CYP2E1 are associated with an increased susceptibility to chemical toxicity and carcinogenesis. This study investigated the induction of hepatic CYP2E1 by ethanol and nicotine, alone and in combination, in a nonhuman primate model. Monkeys that self-administered ethanol and that received subcutaneous injections of nicotine (0.5 mg/kg b.i.d.), alone and in combination, were compared with control animals (four groups, n = 10/group). Chlorzoxazone (CZN) was used as a probe drug to phenotype in vivo CYP2E1 activity before and after chronic ethanol and/or nicotine exposure. CYP2E1 protein levels and in vitro chlorzoxazone metabolism were assessed in liver microsomes. Average daily ethanol consumption was ≈3.0 g/kg (blood ethanol levels ≈24 mM) and was unaffected by nicotine treatment. Ethanol self-administration and nicotine treatment, alone and in combination, significantly increased in vivo CZN disposition compared with that in control animals. The effect of ethanol was only observed at higher levels of intake. Ethanol and nicotine increased CYP2E1 protein levels and in vitro CZN metabolism, with combined exposure to both drugs resulting in the greatest increase. The effect of ethanol was also dependent on level of intake. Chronic exposure to ethanol and nicotine induced hepatic CYP2E1 activity and protein levels, particularly when both drugs were used in combination and when ethanol intake was high. These results have important implications for public health, given the association between elevated CYP2E1 and disease, and the large proportion of individuals who are exposed to ethanol and nicotine, often in combination.

Nicotine induces brain CYP enzymes: relevance to Parkinson's disease.

Brain expression of cytochromes P450 2B6, 2D6 and 2E1 is higher in smokers, and is induced by nicotine in animals. These enzymes can metabolize many of the neurotoxins associated with Parkinson's disease. Since smoking is known to be protective against Parkinson's disease, we hypothesise that nicotine-induced elevation of brain CYPs in smokers may contribute to neuroprotection against Parkinson's disease. This supports the therapeutic use of nicotine to delay the progress of this disease.

Regional and cellular distribution of CYP2D subfamily members in rat brain.

1. Human CYP2D6 is present in brain, metabolizes many drugs and has been implicated in Parkinson's and Alzheimer's diseases and some cancers. It is still unclear which of the six known rat CYP2D subfamily members is/are homologous to human CYP2D6. 2. In this study, RT-PCR, Southern and Western blotting and immunohistochemical techniques were used to study the distribution of CYP2D subfamily member mRNA and proteins across 10 rat brain regions. CYP2D subfamily mRNA and protein levels were correlated with brain dextromethorphan O-demethylation (DOD), a measure of human CYP2D6 and rat CYP2D1 activities. 3. The data showed a strong relationship between CYP2D1 and CYP2D1-18 with brain DOD activity. In addition, it was shown that CYP2D proteins are present in brain mitochondrial as well as microsomal membranes. CYP2D subfamily member mRNA and proteins varied across brain regions and were highly concentrated in specific cell types. 4. These data strongly suggest that CYP2D1 and not CYP2D5 mediates DOD activity in rat brain, and may be the rat homologue of human CYP2D6. The highly localized nature of CYP2D indicates that in specific neurones enzyme levels may approach hepatic levels and, hence, contribute to local alterations in brain drug metabolism.

Regional and cellular induction of nicotine-metabolizing CYP2B1 in rat brain by chronic nicotine treatment.

In the rat, nicotine is metabolized to cotinine primarily by hepatic cytochrome P450 (CYP) 2B1. This enzyme is also found in other organs such as the lung and the brain. Hepatic nicotine metabolism is unaltered after nicotine exposure; however, nicotine may regulate CYP2B1 in other tissues. We hypothesized that nicotine induces its own metabolism in brain by increasing CYP2B1. Male rats were treated with nicotine (0.0, 0.1, 0.3, or 1.0 mg base/kg in saline) s.c. daily for 7 days. CYP2B1 mRNA and protein were assayed in the brain and liver by reverse transcriptase-polymerase chain reaction (RT-PCR), immunoblotting, and immunocytochemistry. In control rats, CYP2B1 mRNA and protein expression were brain region- and cell-specific. CYP2B1 was not induced in the liver, but CYP2B1 mRNA and protein showed dose-dependent, region- and cell-specific patterns of induction across brain regions. At 1.0 mg nicotine/kg, the largest increase in protein was in the brain stem (5.8-fold, P < 0.05) with a corresponding increase in CYP2B1 mRNA (7.6-fold, P < 0.05). Induction of CYP2B1 was also observed in the frontal cortex, striatum, and olfactory tubercle. Immunocytochemistry showed that induction was restricted principally to neurons. These data indicate that nicotine may alter its own metabolism in the brain through transcriptional regulation, perhaps contributing to central tolerance to the effects of nicotine. CYP2B1 and its human homologue CYP2B6 also activate tobacco smoke procarcinogens such as NNK [4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone]. Highly localized increases in CYP2B could result in increased mutagenesis. These data suggest roles for nicotine-induced CYP2B in central metabolic tolerance, nicotine-induced neurotoxicity, neuroplasticity, and carcinogenesis.

The distribution of a CRF-like diuretic peptide in the blood-feeding bug Rhodnius prolixus.

The blood-feeding bug Rhodnius prolixus ingests a large blood meal, and this is followed by a rapid diuresis to eliminate excess water and salt. Previous studies have demonstrated that serotonin and an unidentified peptide act as diuretic factors. In other insects, members of the corticotropin-releasing factor (CRF)-related peptide family have been shown to play a role in post-feeding diuresis. Using fluorescence immunohistochemistry and immunogold labelling with antibodies to the Locusta CRF-like diuretic hormone (Locusta-DH) and serotonin, we have mapped the distribution of neurones displaying these phenotypes in R. prolixus. Strong Locusta-DH-like immunoreactivity was found in numerous neurones of the central nervous system (CNS) and, in particular, in medial neurosecretory cells of the brain and in posterior lateral neurosecretory cells of the mesothoracic ganglionic mass (MTGM). Positively stained neurohaemal areas were found associated with the corpus cardiacum (CC) and on abdominal nerves 1 and 2. In addition, Locusta-DH-like immunoreactive nerve processes were found over the posterior midgut and hindgut. Double-labelling studies for Locusta-DH-like and serotonin-like immunoreactivity demonstrated some co-localisation in the CNS; however, no co-localisation was found in the medial neurosecretory cells of the brain, the posterior lateral neurosecretory cells of the MTGM or neurohaemal areas. To confirm the presence of a diuretic factor in the CC and abdominal nerves, extracts were tested in Malpighian tubule secretion assays and cyclic AMP assays. Extracts of the CC and abdominal nerves caused an increase in the rate of secretion and an increase in the level of cyclic AMP in the Malpighian tubules of fifth-instar R. prolixus. The presence of the peptide in neurohaemal terminals of the CC and abdominal nerves that are distinct from serotonin-containing terminals indicates that the peptide is capable of being released into the haemolymph and that this release can be independent of the release of serotonin.

Characterization of cytochrome P-450 2D1 activity in rat brain: high-affinity kinetics for dextromethorphan.

We investigated the enzymatic function, stability, and regional distribution of rat brain cytochrome P-450 (CYP) 2D1 activity. CYP2D1 is the homolog of human CYP2D6, a genetically variable enzyme that activates or inactivates many clinical drugs acting on the central nervous system (e.g., antidepressants, monoamine oxidase inhibitors, serotonin uptake inhibitors, and neuroleptics), drugs of abuse (e.g., amphetamine and codeine), neurotoxins (e.g., 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, 1,2,3, 4-tetrahydroquinoline), and endogenous neurochemicals (e.g., tryptamine). The CYP2D family has been identified in rodent, canine, and primate brain. Conversion of dextromethorphan to dextrorphan by rat brain membranes was assayed by HPLC and was dependent on NADPH, protein concentration, and incubation time. Significant loss of activity was observed in some homogenizing buffers and after freezing of whole tissues or membrane preparations. Dextromethorphan (0.5-640 microM) metabolism was mediated by high- and low-affinity enzyme systems; K(m1) was 2.7 +/- 2.6 and K(m2) was 757 +/- 156 microM (n = 3 rats, mean +/- S.E.). The enzyme activity was significantly (p <.01) and stereoselectively inhibited by CYP2D1 inhibitors quinine and quinidine (not by CYP2C or CYP3A inhibitors), and by anti-CYP2D6 peptide antiserum (not by anti-CYP2C, -CYP2B, or -CYP3A antibodies). The enzymatic activity demonstrated significant brain regional variation (n = 10 regions, p <.001). These data characterize CYP2D1-mediated dextromethorphan metabolism in rat brain and suggest that localized metabolism of other CYP2D1 substrates (drugs, neurotoxins, and possibly endogenous compounds) within the brain will occur. In humans, CYP2D6 is genetically polymorphic; the variable expression of brain CYP2D6 may result in interindividual differences in central drug and neurotoxin metabolism, possibly contributing to interindividual differences in drug effects and neurotoxicity.

Localization and neurohemal release of FMRFamide-related peptides in the stick insect Carausius morosus.

FMRFamide-like immunoreactivity was localized immunohistochemically in the central stomatogastric nervous systems, visceral tissues, and the neurohemal corpora cardiaca, transverse, and segmental nerves. Each of these neurohemal areas contains one morphologically distinct type of immunoreactive neurosecretory granule. The hemolymph level of FMRFamide-like peptides, quantified by RIA, is higher in animals sampled 2 h into the dark cycle, relative to those sampled at mid-light cycle or 9 h into the dark cycle. High potassium depolarization evokes the calcium-dependent release of FMRFamide-like peptides from neurohemal areas in vitro and HPLC fractionation of hemolymph, corpora cardiaca, and their bathing medium suggests that these organs contribute a single peptide to the FMRFamide-related peptides circulating in the hemolymph of active animals.

Immunogold labelling of serotonin-like and FMRFamide-like immunoreactive material in neurohaemal areas on abdominal nerves of Rhodnius prolixus.

The ultrastructure of neurohaemal areas on abdominal nerves of the blood-sucking bug Rhodnius prolixus was investigated. Four types of axon terminals were found, distinguished by the morphology of their neurosecretory granules. By use of post-embedding immunogold labelling, granules in Type I axon terminals were shown to contain serotonin-like immunoreactive material, and granules in Type II axon terminals were shown to contain FMRFamide-like immunoreactive material. There was no colocalization of these materials. It is suggested that Type III terminals contain peptidergic diuretic hormone, which has previously been reported to be present in electron-dense neurosecretory granules in this neurohaemal area. The identity of material in Type IV terminals is unknown.

The effect of the brain and dorsal bodies of Helisoma duryi (mollusca: pulmonata) on albumen gland synthetic activity in vitro.

Albumen gland explants from Helisoma duryi can be maintained in a liquid culture medium for at least 96 hr. This allows measurement of incorporation of [14C]glucose into newly synthesized polysaccharides both retained in the tissue and released into the medium. The albumen gland basal synthetic activity is much higher in reproductively active than in reproductively inactive snails (maintained in constant darkness, virgin, or without dorsal bodies). The presence of the whole brain or cerebral ganglia (with dorsal bodies) of reproductively active snails in the culture medium stimulates polysaccharide synthesis in albumen gland explants from snails in constant darkness. Addition of dorsal body homogenate to the culture medium stimulates polysaccharide synthesis in albumen gland explants from snails without dorsal bodies. These results show that an endocrine factor from the dorsal bodies and possibly a neuroendocrine factor(s) from the cerebral ganglia directly stimulate polysaccharide synthesis in the albumen gland.