Stress-Induced Changes in Nucleocytoplasmic Localization of Crucial Factors in Gene Expression Regulation.

This study investigates the toxic effect of harmful materials, unfiltered by the placenta, on neonatal umbilical cord (UC) vessels, focusing on stress-induced adaptations in transcriptional and translational processes. It aims to analyze changes in pathways related to mRNA condensate formation, transcriptional regulation, and DNA damage response under maternal smoking-induced stress. UC vessels from neonates born to smoking (Sm) and nonsmoking mothers (Ctr) were examined. Immunofluorescence staining and confocal microscopy assessed the localization of key markers, including Transcription Complex Subunit 1 (CNOT1) and the largest subunit of RNA polymerase II enzyme (RPB1). Additionally, markers of DNA damage response, such as Poly(ADP-ribose) polymerase-1, were evaluated. In Sm samples, dissolution of CNOT1 granules in UC vessels was observed, potentially aiding stalled translation and enhancing transcription via RPB1 assembly and translocation. Control vessels showed predominant cytoplasmic RPB1 localization. Despite adaptive responses, Sm endothelial cells exhibited significant damage, indicated by markers like Poly(ADP-ribose) polymerase-1. Ex vivo metal treatment on control vessels mirrored Sm sample alterations, emphasizing marker roles in cell survival under toxic exposure. Maternal smoking induces specific molecular adaptations in UC vessels, affecting mRNA condensate formation, transcriptional regulation, and DNA damage response pathways. Understanding these intricate molecular mechanisms could inform interventions to improve neonatal health outcomes and mitigate adverse effects of toxic exposure during pregnancy.

Molecular Background of Toxic-Substances-Induced Morphological Alterations in the Umbilical Cord Vessels and Fetal Red Blood Cells.

The relationship between smoking and human health has been investigated mostly in adults, despite the fact that the chemicals originating from sustained maternal smoking disrupt the carefully orchestrated regulatory cascades in the developing fetus. In this study, we followed molecular alterations in the umbilical cord (UC) vessels and fetal red blood cells (RBCs), which faithfully reflect the in vivo status of the fetus. We showed evidence for the decreased level of DNA-PKcs-positive nuclei in samples with smoking origin, which is associated with the impaired DNA repair system. Furthermore, we pointed out the altered ratio of MMP-9 metalloproteinase and its endogenous inhibitor TIMP-1, which might be a possible explanation for the morphological abnormalities in the UC vessels. The presented in vivo dataset emphasizes the higher vulnerability of the veins, as the primary target for the toxic materials unfiltered by the placenta. All these events become amplified by the functionally impaired fetal RBC population via a crosstalk mechanism between the vessel endothelium and the circulating RBCs. In our ex vivo approach, we looked for the molecular explanation of metal-exposure-induced alterations, where expressions of the selected genes were upregulated in the control group, while samples with smoking origin showed a lack of response, indicative of prior long-term in utero exposure.

Protective effect of green tea against neuro-functional alterations in rats treated with MnO2 nanoparticles.

BACKGROUND: Inhalation of manganese-containing metal fumes at workplaces can cause central nervous damage including a Parkinson-like syndrome. Oxidative stress is likely to be involved in the pathomechanism, due to the presence of nano-sized metal oxide particles with high biological and chemical activity. Oxidative damage of the nervous system could be prevented or ameliorated by properly applied antioxidants, preferably natural ones such as green tea, a popular drink. The aim of this work was to see if orally applied green tea brew could diminish the functional neurotoxicity of manganese dioxide nanoparticles introduced into the airways of rats. RESULTS: Young adult male Wistar rats were treated intratracheally for 6 weeks with a suspension of synthetic MnO2 nanoparticles (4 mg/kg body weight), and received green tea brew (1 g leaves 200 mL-1 water) as drinking fluid. Reduced body weight gain, indicating general toxicity of the nanoparticles, was not influenced by green tea. However, in rats receiving green tea the nervous system effects - changes in the spontaneous and evoked cortical activity and peripheral nerve action potential - were diminished. CONCLUSION: The use of green tea as a neuroprotective functional drink seems to be a viable approach. © 2016 Society of Chemical Industry.

Elevated levels of macromolecular damage are correlated with increased nitric oxide synthase expression in erythrocytes isolated from twin neonates.

Pregnancy is a state associated with an enhanced metabolism and demand for O2 , which may lead to the overproduction of reactive oxygen species (ROS) and hence to oxidative stress. An elevated ROS level may result in delayed development and a low birth weight. The aim of this study was to reveal the consequences of multiple pregnancies on the redox status of neonatal human red blood cells (RBCs) and evaluate the role of endothelial nitric oxide synthase (NOS3) - expressing RBCs in the generation of oxidative stress. The study presents evidence of higher levels of production of hydrogen peroxide, peroxynitrite and nitrate content in the RBCs of twin neonates, clearly reflected by an elevated level of protein and lipid damages. This phenotype appears to be a consequence of multiple pregnancies, regardless of the level of maturity or the birth weight of the twins. Besides the higher level of ROS, there was a general decrease in the expression of genes coding for antioxidants. The first data are presented on NOS3-expressing neonatal human RBCs. The number of RBCs producing NOS3 was more than twice as high in twin neonates compared to singletons, with no correlation to maturity.

Green tea and vitamin C ameliorate some neuro-functional and biochemical signs of arsenic toxicity in rats.

BACKGROUND/OBJECTIVES: Nervous system damage is one of the consequences of oral exposure to waterborne inorganic arsenic. In this work, the role of oxidative status in the neurotoxicity of arsenic and the possible role of two foodborne antioxidants in ameliorating arsenic-related oxidative stress were investigated. METHODS: Male Wistar rats were given 10 mg/kg b.w. of trivalent inorganic arsenic (in the form of NaAsO2), 5 day/week for 6 weeks by gavage, combined with vitamin C solution (1 g/l) or green tea infusion (2.5 g in 500 ml boiled water) as antioxidants given in the drinking fluid. RESULTS: Body weight gain was reduced by arsenic from the second week and the antioxidants had no effect on that. Cortical evoked potentials had increased latency, tail nerve conduction velocity was reduced, and this latter effect was counteracted by the antioxidants. The effect of green tea was stronger than that of vitamin C, and green tea also diminished lipid peroxidation induced by As. There was fair correlation between brain As levels, electrophysiological changes, and lipid peroxidation, suggesting a causal relationship. DISCUSSION: Natural antioxidants might be useful in the protection of the central nervous system against the toxicity of oral As.

Ldb1 is essential for development of Nkx2.1 lineage derived GABAergic and cholinergic neurons in the telencephalon.

The progenitor zones of the embryonic mouse ventral telencephalon give rise to GABAergic and cholinergic neurons. We have shown previously that two LIM-homeodomain (LIM-HD) transcription factors, Lhx6 and Lhx8, that are downstream of Nkx2.1, are critical for the development of telencephalic GABAergic and cholinergic neurons. Here we investigate the role of Ldb1, a nuclear protein that binds directly to all LIM-HD factors, in the development of these ventral telencephalon derived neurons. We show that Ldb1 is expressed in the Nkx2.1 cell lineage during embryonic development and in mature neurons. Conditional deletion of Ldb1 causes defects in the expression of a series of genes in the ventral telencephalon and severe impairment in the tangential migration of cortical interneurons from the ventral telencephalon. Similar to the phenotypes observed in Lhx6 or Lhx8 mutant mice, the Ldb1 conditional mutants show a reduction in the number of both GABAergic and cholinergic neurons in the telencephalon. Furthermore, our analysis reveals defects in the development of the parvalbumin-positive neurons in the globus pallidus and striatum of the Ldb1 mutants. These results provide evidence that Ldb1 plays an essential role as a transcription co-regulator of Lhx6 and Lhx8 in the control of mammalian telencephalon development.

Impact of acute arsenic and cadmium exposure on the expression of two haeme oxygenase genes and other antioxidant markers in common carp (Cyprinus carpio).

The aim was to study the effects of cadmium (Cd) and arsenic (As) on haeme oxygenases (HOs) and other oxidative stress biomarkers, and their roles in macromolecule damage in liver and kidney of common carp (Cyprinus carpio L.). HOs play a critical role in the defence system against oxidative damage, producing biliverdin and carbon monoxide with important free radical scavenging properties. However, increased HO activity in haeme degradation may also lead to a pro-oxidant effect through the liberation of Fe-modifying Cd and As toxicity. The response of an organism to exposure to toxic metals is in many cases brought about by changes at the level of gene expression. In this study, the genes ho-1 and ho-2 of the common carp were identified, and the changes in gene expressions were analysed from the aspect of Cd and As accumulation. Both ho-1 and ho-2 are transcriptionally induced by Cd and As, but their inductions differ in time course, dose response and tissue specificity. The expression of ho1 was mostly affected by As, primarily in the liver (45-fold), whereas it was enhanced with higher efficacy by Cd in the kidney (25-fold). The cellular redox status and the damage of lipid molecules were monitored via the ratio of reduced to oxidized glutathione, the levels of H2 O2 and lipid peroxidation, and the activities of superoxide dismutase (SOD) and catalase (CAT).

Molecular Study on Twin Cohort with Discordant Birth Weight.

The increased rate of twinning has pointed out newer challenges in clinical practices related to gestational complications, intrauterine growth restriction, perinatal mortality, and comorbidities. As a twin pregnancy progresses, the increased demand for oxygen supply can easily disrupt the redox homeostasis balance and further impose a greater challenge for the developing fetuses. A substantial birth-weight difference acts as an indicator of a deficit in oxygenation or blood flow to one of the fetuses, which might be related to a low bioavailable nitric oxide level. Therefore, in this study, we focused on networks involved in the adjustment of oxygen supply, like the activation of inducible and endothelial nitric oxide synthase (NOS3) along with free radical and lipid peroxide formation in mature twin pairs with high birth-weight differences. The selected parameters were followed by immunofluorescence staining, fluorescence-activated cell sorting analysis, and biochemical measurements in the umbilical cord vessels and fetal red blood cells. Based on our data set, it is clear that the lower-weight siblings are markedly exposed to persistent intrauterine hypoxic conditions, which are connected to a decreased level in NOS3 activation. Furthermore, the increased level of peroxynitrite aggravates lipid peroxidation and induces morphological and functional damage and loss in redox homeostasis.

Gut region-specific diabetic damage to the capillary endothelium adjacent to the myenteric plexus.

OBJECTIVE: Damage in the capillaries supplying the MP has been proposed as a critical factor in the development of diabetic enteric neuropathy. We therefore investigated connections between STZ-induced diabetes and the BM morphology, the size of caveolar compartments, the width of TJs, the transport of albumin, and the quantitative features of Cav-1 and eNOS expression in these microvessels. METHODS: Gut segments from diabetic rats were compared with those from insulin-treated diabetics and those from controls. The effects of diabetes on the BM, the caveolar compartments, and the TJs were evaluated morphometrically. The quantitative features of the albumin transport were investigated by postembedding immunohistochemistry. The diabetes-related changes in Cav-1 and eNOS expression were assessed by postembedding immunohistochemistry and molecular method. RESULTS: Thickening of the BM, enlargement of the caveolar compartments, opening of the junctions, enhanced transport of albumin, and overexpression of Cav-1 and eNOS were documented in diabetic animals. Insulin replacement in certain gut segments prevented the development of these alterations. CONCLUSIONS: These data provide morphological, functional, and molecular evidence that the endothelial cells in capillaries adjacent to the MP is a target of diabetic damage in a regional manner.

A role of the LIM-homeobox gene Lhx2 in the regulation of pituitary development.

The mammalian pituitary gland originates from two separate germinal tissues during embryonic development. The anterior and intermediate lobes of the pituitary are derived from Rathke's pouch, a pocket formed by an invagination of the oral ectoderm. The posterior lobe is derived from the infundibulum, which is formed by evagination of the neuroectoderm in the ventral diencephalon. Previous studies have shown that development of Rathke's pouch and the generation of distinct populations of hormone-producing endocrine cell lineages in the anterior/intermediate pituitary lobes is regulated by a number of transcription factors expressed in the pouch and by inductive signals from the ventral diencephalon/infundibulum. However, little is known about factors that regulate the development of the posterior pituitary lobe. In this study, we show that the LIM-homeobox gene Lhx2 is extensively expressed in the developing ventral diencephalon, including the infundibulum and the posterior lobe of the pituitary. Deletion of Lhx2 gene results in persistent cell proliferation, a complete failure of evagination of the neuroectoderm in the ventral diencephalon, and defects in the formation of the distinct morphological features of the infundibulum and the posterior pituitary lobe. Rathke's pouch is formed and endocrine cell lineages are generated in the anterior/intermediate pituitary lobes of the Lhx2 mutant. However, the shape and organization of the pouch and the anterior/intermediate pituitary lobes are severely altered due to the defects in development of the infundibulum and the posterior lobe. Our study thus reveals an essential role for Lhx2 in the regulation of posterior pituitary development and suggests a mechanism whereby development of the posterior lobe may affect the development of the anterior and intermediate lobes of the pituitary gland.

Fetal oxygen supply can be improved by an effective cross-talk between fetal erythrocytes and vascular endothelium.

In twin/multiple pregnancy, siblings experience an adverse intrauterine environment which forms the major etiological factor leading to pathological conditions. The status of the developing fetus is highly determined by the nitric oxide (NO) level, that facilitates vasodilation which in turn modulates the oxygen and nutrition supply. As the umbilical cord (UC) lacks innervation, activation of the endothelial nitric oxide synthase (NOS3) is fundamental to maintain adequate NO production. Recent ground breaking fact showed that under stress conditions, circulating red blood cells (RBCs) can actively produces NO as a "rescue mechanism". Therefore, this study majorly focused on the molecular mechanisms that affected the redox environment by altering NOS3 activation - both in the UC arteries and vein endothelium and RBCs - that have impacts on developmental parameters, like birth weight. In connection to that, we pursued the communication efficiency between the vessels' endothelium and the circulating RBCs in demand of bioavailable NO. Our results indicated that twinning itself at stage 33-35 weeks, does not reduce the NOS3 level and its phosphorylation status in the cord vessels. However, RBC-NOS3 activation is highly upregulated during this period - providing additional evidence for the active regulatory role of fetal RBCs in the rate of blood flow - and this functional activity highly correlates with the birth weight of the fetuses. Detailed analysis on NOS3 signalling at different time points of gestation could establish a benchmark in understanding of the pathophysiological mechanisms involved in the process of developing neonatal vascular diseases.

Diabetes-related intestinal region-specific thickening of ganglionic basement membrane and regionally decreased matrix metalloproteinase 9 expression in myenteric ganglia.

BACKGROUND: The importance of the neuronal microenvironment has been recently highlighted in gut region-specific diabetic enteric neuropathy. Regionally distinct thickening of endothelial basement membrane (BM) of intestinal capillaries supplying the myenteric ganglia coincide with neuronal damage in different intestinal segments. Accelerated synthesis of matrix molecules and reduced degradation of matrix components may also contribute to the imbalance of extracellular matrix dynamics resulting in BM thickening. Among the matrix degrading proteinases, matrix metalloproteinase 9 (MMP9) and its tissue inhibitor (TIMP1) are essential in regulating extracellular matrix remodelling. AIM: To evaluate the intestinal segment-specific effects of diabetes and insulin replacement on ganglionic BM thickness, MMP9 and TIMP1 expression. METHODS: Ten weeks after the onset of hyperglycaemia gut segments were taken from the duodenum and ileum of streptozotocin-induced diabetic, insulin-treated diabetic and sex- and age-matched control rats. The thickness of BM surrounding myenteric ganglia was measured by electron microscopic morphometry. Whole-mount preparations of myenteric plexus were prepared from the different gut regions for MMP9/TIMP1 double-labelling fluorescent immunohistochemistry. Post-embedding immunogold electron microscopy was applied on ultrathin sections to evaluate the MMP9 and TIMP1 expression in myenteric ganglia and their microenvironment from different gut segments and conditions. The MMP9 and TIMP1 messenger ribonucleic acid (mRNA) level was measured by quantitative polymerase chain reaction. RESULTS: Ten weeks after the onset of hyperglycaemia, the ganglionic BM was significantly thickened in the diabetic ileum, while it remained intact in the duodenum. The immediate insulin treatment prevented the diabetes-related thickening of the BM surrounding the ileal myenteric ganglia. Quantification of particle density showed an increasing tendency for MMP9 and a decreasing tendency for TIMP1 from the proximal to the distal small intestine under control conditions. In the diabetic ileum, the number of MMP9-indicating gold particles decreased in myenteric ganglia, endothelial cells of capillaries and intestinal smooth muscle cells, however, it remained unchanged in all duodenal compartments. The MMP9/TIMP1 ratio was also decreased in ileal ganglia only. However, a marked segment-specific induction was revealed in MMP9 and TIMP1 at the mRNA levels. CONCLUSION: These findings support that the regional decrease in MMP9 expression in myenteric ganglia and their microenvironment may contribute to extracellular matrix accumulation, resulting in a region-specific thickening of ganglionic BM.

Sustained Maternal Smoking Triggers Endothelial-Mediated Oxidative Stress in the Umbilical Cord Vessels, Resulting in Vascular Dysfunction.

Nitric oxide (NO) bioavailability is fundamental in the regulation of redox balance and functionality of the endothelium, especially in the case of the umbilical cord (UC), which has no innervation. The analysis of UC vessel-related complications could serve as a useful tool in the understanding of the pathophysiological mechanisms leading to neonatal cardiovascular disorders. Therefore, the aim of this study was to characterize the mechanisms that rule the severity of prenatal endothelial dysfunction, induced by the long-term effect of maternal smoking. Our analysis describes the initiation and the consequences of endothelial nitric oxide synthase (NOS3) deactivation, along with the up-regulation of possible compensatory pathways, using structural, molecular and biochemical approaches. This study was carried out on both the UC arteries and veins originated from neonates born to non-smoking and heavy-smoking mothers. The alterations stimulated by maternal smoking are vessel-specific and proportional to the level of exposure to harmful materials passed through the placenta. Typically, in the primarily exposed veins, an increased formation of reactive oxygen species and an up-regulation of the highly-efficient NOS2-NO producing pathway were detected. Despite all the extensive structural and functional damages, the ex vivo heat and cadmium ion-treated UC vein pieces still support the potential for stress response.

Erratum to "Maternal Smoking Highly Affects the Function, Membrane Integrity, and Rheological Properties in Fetal Red Blood Cells".

[This corrects the article DOI: 10.1155/2019/1509798.].

Expressions of heat shock and metallothionein genes in the heart of common carp (Cyprinus carpio): effects of temperature shock and heavy metal exposure.

Heat shock proteins (HSPs) and metallothioneins (MTs) play important roles in protection against environmental stressors. The present study analyzes and compares the regulation of heat shock ( hsp70, hsc70-1 and hsp90alpha ) and metallothionein (MT-1 and MT-2) genes in the heart of common carp, in response to elevated temperature, cold shock and exposure to several heavy metal ions (As 3+ , Cd 2+ and Cu 2+ ), in whole-animal experiments. Among these metals, arsenate proved to be the most potent inducer of the examined stress genes; the hsp90alpha and MT-1 mRNA levels were elevated 11- and 10-fold, respectively, after a 24-h exposure. In contrast, Cd 2+ at 10 mg/L had no impact on the expression of hsp90alpha , and the MT genes also proved to be rather insensitive to Cd 2+ treatment in the heart: only a 2-2.5-fold induction was observed in response to 10 mg/L Cd 2+ . Heat shock resulted in a transient induction of hsp70 (19-fold) and hsp90alpha (15-fold), while elevated temperature had no effect on the expression of the MTs. Direct cold shock induced hsp70 expression (14-fold), while the hsp90alpha (26-fold) and MT-2 (2-fold) expressions peaked after the recovery period following a direct cold shock. The five stress genes examined in this study exhibited a unique, tissue-specific basal expression pattern and a characteristic sensitivity to metal treatments and temperature shocks.

Mature Twin Neonates Exhibit Oxidative Stress via Nitric Oxide Synthase Dysfunctionality: A Prognostic Stress Marker in the Red Blood Cells and Umbilical Cord Vessels.

Intrauterine hypoxic condition increases the generation of reactive oxygen species and fetal oxidative stress. Multiple pregnancy always bears an additional oxidative stress condition with severe complications, such as prematurity, structural abnormalities, delayed development and low birthweight. The umbilical cord (UC) vessels, along with circulating fetal red blood cells (RBCs), highly determine the oxygenation status of fetus and regulate the feto-placental circulation. As UC lacks innervation, the activation of the endothelial nitric oxide synthase (NOS3) is fundamental for proper NO production. Therefore, we aimed to study the NOS3 activation pathways along with damages to macromolecules in the endothelium of UC vessels and RBCs of mature non-discordant twins, in connection to major differences in their birth weight. We provide evidence that, under severe hypoxic conditions such as twin pregnancy, the NOS3-related NO production pathways are altered both in UC vessels and RBCs; moreover, the extent of changes is highly birthweight-specific. Furthermore, macromolecular damages are prominent in the RBCs and arteries compared to the vein, with a similar increase in the Arginase1 level, which is believed to play a role in NOS3 functionality, resulting in endothelial dysfunctionality, which might have relevance to the major etiologies of cardiovascular diseases in later life.

Response of Human Neutrophil Granulocytes to the Hyphae of the Emerging Fungal Pathogen Curvularia lunata.

Curvularia lunata is an ascomycete filamentous fungus causing local and invasive phaeohyphomycoses in both immunocompromised and immunocompetent patients. Neutrophils are crucial participants of the first line host defense against fungal infections. They migrate to the infected site and eliminate the infectious agents by various mechanisms including phagocytoses, oxidative damage, or formation of neutrophil extracellular trap (NET). Neutropenia may be a risk factor for phaeohyphomycoses, and restoration of the neutrophil function can improve the outcome of the infection. In the present study, interaction of primary human neutrophil granulocytes with the hyphae C. lunata was examined and compared to that with the well characterized filamentous fungal pathogen Aspergillus fumigatus. Neutrophils could recognize the serum opsonized hyphae of C. lunata and attach to them. Myeloperoxidase release was also activated by a soluble factor present in the culture supernatant of the fungus. Induction of the oxidative burst was found to depend on serum opsonization of the hyphae. Although extracellular hydrogen peroxide production was induced, the fungus efficiently blocked the oxidative burst by acidifying the reaction environment. This blockage also affected the NET forming ability of the neutrophils.

Sustained maternal smoking-associated changes in the physico-chemical properties of fetal RBC membranes might serve as early markers for vascular comorbidities.

Maternal smoking-induced congenital heart and microvascular defects are closely associated with the impaired functioning of the in-utero feto-placental circulation system. Current groundbreaking facts revealed intimate crosstalk between circulating red blood cells (RBCs) and the vascular endothelium. Thus, RBCs have become the protagonists under varied pathological and adverse pro-oxidative cellular stress conditions. We isolated and screened fetal RBCs from the arterial cord blood of neonates, born to non-smoking (RBC-NS) and smoking mothers (RBC-S), assuming that parameters of fetal RBCs are blueprints of conditions experienced in-utero. Using atomic force microscopy and mass spectrometry-based shotgun lipidomics in the RBC-S population we revealed induced membrane stiffness, loss in intrinsic plastic activities and several abnormalities in their membrane-lipid composition, that could consequently result in perturbed hemodynamic flow movements. Altogether, these features are indicative of the outcome of neonatal microvascular complications and suggest unavailability for the potential rescue mechanism in cases of vascular endothelium impairment due to altered membrane integrity and rheological properties.

Endothelial nitric oxide synthase is up-regulated in the umbilical cord in pregnancies complicated with intrauterine growth retardation.

BACKGROUND: NO and NO synthases (NOS) play an important role in the physiology of the fetomaternal blood circulation, although their expression in pathological conditions is unclear. Intrauterine growth retardation (IUGR) is a disorder most probably caused by abnormality of the fetomaternal bloodflow. MATERIALS AND METHODS: The expression of endothelial NOS (ecNOS) from arteria umbilicalis and the nitrite and peroxynitrite level of umbilical blood were determined. Major consequences of peroxynitrite toxicity are lipid peroxidation and glutathione depletion; these parameters were also measured. Finally, superoxide dismutase (SOD) activity was assayed to evaluate the level of superoxide anions. RESULTS: Elevated expression of ecNOS was found to be coupled with significantly lower SOD activity and glutathione level, and increased lipid peroxidation in IUGR neonates. CONCLUSION: The increased NO indices could represent a compensatory effort to improve placental bloodflow, but in IUGR neonates it is coupled with inadequate antioxidant defence, resulting in significant oxidative stress.

Expression of two metallothionein genes in different brain regions of common carp.

The expression pattern of two metallothionein (MT) genes in response to temperature shock and exposure to Cd(2+) was investigated in the brain of common carp ( Cyprinus carpio ), in whole-animal experiments. The changes in the levels of MT-1 and MT-2 mRNA in the olfactory lobe, midbrain and cerebellum were followed by semiquantitative RT-PCR. The inducibility of the two MT genes was brain region and stressor-specific. Cd(2+) affected mostly the expression of MT-2, while the level of the MT-1 transcript did not change significantly in any of the brain regions examined. Moreover, the MT-2 expression was regulated spatially; MT-2 was induced significantly more strongly in the olfactory lobe than in the cerebellum or midbrain. A sudden temperature drop mainly affected the expression of the MT-1 gene; after 5 h of cold shock, the MT-1 mRNA level was about 25% of the basal value in the cerebellum and the midbrain region. The MT-2 expression did not change significantly during this treatment.