Search

Messenger RNA, obtained from post-mortem brain of 10 schizophrenics, five depressed patients and 10 control subjects, was characterised with respect to a number of parameters. It was found that post-mortem delay was not the major factor in determining RNA yield, size (as determined by cDNA synthesis) and biological activity. Biological activity, as determined by in vitro translation in a reticulocyte-lysate system, could be observed using messenger RNA from periods of 0 to 84 hours post-mortem. Two-dimensional gel analysis of the newly-synthesised radiolabelled products obtained from this material revealed several hundred individual species but no consistent degradation of any particular species with post-mortem delay. It is suggested, therefore, that premortem changes are as important as post-mortem changes in determining RNA yield, size and biological activity. Although no consistent difference could be found between patients and controls using any of these parameters, this study confirms that, by isolating messenger RNA from post-mortem human brain, valuable information can be gained on gene expression in psychiatric disorders. [ABSTRACT FROM AUTHOR]

Erythrocytes from lithium treated patients were separated according to the time they had been circulating in the plasma. Choline transport and choline content were measured in the erythrocyte fractions separated according to age, in order to investigate the relationship between decreased choline transport and increased choline content seen in lithium treated patients. The most recently formed erythrocytes of normal subjects had the greatest choline content and the most active choline carrier. The erythrocytes of lithium treated patients had reduced choline carrier activity and increased choline content in all age bands. However the greatest accumulation of choline and least inhibited choline carrier activity was seen in the most recently formed cells. The alteration in phospholipid concentrations measured could not of themselves account for elevated erythrocyte choline levels seen in lithium treated patients. It is concluded that the increase in choline content levels in lithiated erythrocytes does not have a simple inverse relationship with the deficiency in choline transport. The inhibition of the choline carrier is caused by modification due to circulation in lithiated plasma rather than a lack of its synthesis in reticulocytes. [ABSTRACT FROM AUTHOR]

We have previously reported a changed mitochondrial (mt) gene expression in brain from patients with schizophrenia [Schizophr. Res. 14 (1995) 203]; now, we describe the distribution in the mtDNA from lymphocytes of a heteroplasmic sequence variation that was originally found in the mtDNA from the postmortem brain of a patient with schizophrenia. The variant is m.12027T>C and results in the change from isoleucine to threonine at position 423 of the ND4 subunit of NADH-ubiquinone reductase. Using a PCR-RFLP method, we have determined the heteroplasmy as the ratio of variant to total (variant ratio) at m.12027 in 184 controls and 181 patients with schizophrenia as well as 24 postmortem brain samples. The distribution of variants is bimodal having peaks at variant ratios of 0.262 and 0.732. The variant-rich fraction is very significantly associated with schizophrenia in males (47%), while there is only 18% in control males. There are significantly more variant-rich control females (36%) than control males (18%), suggesting that the female population is less sensitive to the presence of a variant in terms of liability to schizophrenia. In variant-rich samples from postmortem brain originating from both sexes, there is an increased superoxide production, suggesting that the variation contributes to oxidative stress. Antioxidant glycosides, such as quercetin rutoside, quench the superoxide production without (in contrast to neuroleptic drugs) interfering with the electron transfer activity of the reductase.

Basal forebrain tissue fragments taken from embryonic day 15 were separated into primary astrocytes and primary neurons in culture and grafted to rats with alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid lesions to the nucleus basalis and medial septal regions. The two cell types were compared in two experimental paradigms for their behavioural, biochemical and histochemical effects; standard transplants of whole basal forebrain and sham transplants served as positive and negative controls, respectively. Each transplant cell type was characterised by in vitro immunocytochemistry to assess content and purity. Memory deficits produced by the lesions in a spatial win-stay T-maze task (Experiment 1) and a spatial plus associative radial maze task (Experiment 2) were significantly improved by the astrocyte, but not by the neuronal, primary cell transplants. The astrocyte graft groups performed as well as standard cholinergic rich basal forebrain groups, reaching control levels on both tasks, while the neuronal transplant groups were not significantly different to lesioned (sham transplanted) rats. There was no recovery in choline acetyltransferase activity in brain regions containing astrocyte grafts whereas activity in the neuronal graft regions was increased (often to control levels), similar to recovery produced by basal forebrain grafts. Grafts in all groups survived, transplanted neurons displaying similar morphology and placement in the host brain to unseparated basal forebrain grafts, while astrocytes showed evidence of migration. The cultured astrocytes were estimated to be > 95% pure, showing positive staining for all astrocyte markers and an absence of staining for neuronal markers. The results indicate that the restoration of cognitive function following fetal grafting is not dependent upon a restoration of cholinergic neuronal activity but is more likely mediated via diffuse graft-host communication, with trophic secretion a probable factor. This study emphasizes the usefulness of astrocytes in the repair of central nervous system injury and has implications for therapeutic potential.

In order to examine the molecular basis of schizophrenia we have employed a sequential differential hybridisation protocol to isolate mRNAs whose abundances are altered in schizophrenic compared to normal frontal cortex. Five cDNAs present at abnormal levels in the schizophrenic brain have been isolated by this method. The sequences were identified on the basis of homologies in the EMBL and Genbank databases. All the sequences encode mitochondrial transcripts; one encodes part of the 12s rRNA, three encode parts of the 16s rRNA region of the mitochondrial genome whilst the fourth encodes part of the amino acid sequence of cytochrome oxidase sub-unit II. It was established that mitochondrial sequences were not over-represented in the library and that this could therefore not account for the isolation of five mitochondrial transcripts by this procedure. Increased levels of cytochrome oxidase mRNA were detected in a further set of extracts from the frontal cortex of eight schizophrenic patients and five controls. The amount of mt-DNA was measured in these samples but there was no difference between schizophrenic and control. These results indicate a possible abnormality of mitochondrial function in the schizophrenic frontal cortex.

Heterosynaptic short-term depression (STD) of the stratum radiatum and stratum oriens inputs to the CA1 region was studied in rat hippocampal slices. STD was evoked by trains of 1050 impulses with interstimulus interval (ISI) variable from 10 to 700 ms. The STD was found to be very pronounced for tetanizations with ISI around 200 ms, and almost absent for ISI less than 50 ms or more than 500 ms. These data show that theta-like tetanization is an effective pattern not only for induction of the long-term potentiation (LTP), as has been shown previously, but for production of the heterosynaptic STD as well. This implies that heterosynaptic STD can effectively modulate induction of LTP by theta-like tetanization, and plays an important role in differentiation of potentiated pathways. It is discussed that the theta-like tetanization-induced release of ACh is a possible mechanism of the STD.

Rats were tested for spatial performance in a water maze with further in vitro investigation of short-term and long-term potentiation (STP and LTP) in the CA1 region of the hippocampus. Recordings of the population spike in stratum pyramidale and population excitatory postsynaptic potential (EPSP) in stratum radiatum were made with extracellular electrodes after stimulation of the radiatum and oriens inputs in the region. It was found that for both inputs, STP and LTP of the population spike amplitude correlated with behavioral parameters of memory: latency of reaching the hidden platform (escape latency) and percent of time which the animal spent inside the quadrant with the platform. Potentiation of the initial slope of EPSP, in contrast, did not correlate with these parameters. These data support the hypothesis that hippocampal LTP may underlie spatial performance and show that potentiation of the output characteristic of the hippocampus (population spike), but not of the EPSP, is a physiological correlate for spatial memory.

Poly(A+) mRNA was extracted from the post-mortem brain of schizophrenics (9 subjects), unipolar depressives (5 subjects) and controls (10 subjects) and used to direct the in vitro translation of radiolabelled protein in a cell-free reticulocyte-lysate system. Protein species were analysed on two-dimensional gels. Over 200 products were detected and, from these, 74 well-resolved species were chosen for further analysis. The optical density of each product was quantified by image analysis and normalised with respect to overall gel intensity. It was found that 7 novel, uncharacterised protein species, ranging from molecular weights (Mr) 17 kDa to 38 kDa and apparent isoelectric points (pI) 5.7-7.1, changed significantly in intensity in the psychotic groups compared to controls. One species changed only in the schizophrenia group (Mr = 26 kDa, pI = 5.8, 18% of control intensity) and 3 changed only in the depressive group (Mr = 38 kDa, pI = 6.2, 540% of control; Mr = 34 kDa, pI = 6.2, 6% of control; Mr = 17 kDa, pI = 5.7, 238% of control). Three further protein species were common to both psychotic groups (one species decreased in both schizophrenia and depression, Mr = 33 kDa, pI = 5.8; two species showed opposing intensity changes, decreasing in schizophrenia and increasing in depression, Mr = 35 kDa, pI = 7.1; Mr = 23 kDa, pI = 6.1). None of these changes was a function of post-mortem delay or mode of death. It is quite likely that such protein species reflect the abundance of specific mRNAs and target gene systems associated with the disease state.(ABSTRACT TRUNCATED AT 250 WORDS)

Total cellular polyadenylated RNA (poly(A)+ RNA, mRNA) was prepared after guanidinium thiocyanate extraction of frozen brain tissue from age-matched controls and patients suffering from schizophrenia and unipolar depression. These mRNA populations were analysed by in vitro translation followed by two-dimensional gel analysis. Data were obtained from fluorograms derived from 10 different schizophrenic patients, 10 different controls and 5 different depressive patients. The relative concentrations of mRNA species coding for 4 translation products (33 kDa, pI 5.8; 26 kDa, pI 5.8; 35 kDa, pI 7.1; 23 kDa, pI 6.1) were significantly reduced in schizophrenia compared to controls when determined by computerised image analysis of the fluorograms. In the case of depression, the relative concentrations of mRNA species coding for 6 translation products were significantly altered, 4 being increased (38 kDa, pI 6.2, 17 kDa, pI 5.7, 35 kDa, pI 7.1; 23 kDa, pI 6.1) and two decreased (34 kDa, pI 6.2; 33 kDa, pI 5.8). Three translation products were altered in both schizophrenia and depression, one (33 kDa, pI 5.8) being altered according to the same trend, a decrease relative to controls, but two (35 kDa, pI 7.1; 23 kDa, pI 6.1) being altered differently in schizophrenia (reduced) and depression (increased). The effects of post mortem delay, mode of death and drug treatment on mRNA composition were also examined and found not to affect the levels of these translation products significantly. The significance of these changes will be discussed in relation to their relevance of biological mechanisms in the psychoses.

The report of a putative schizophrenia susceptibility gene linked to markers in the chromosome 5q11-q13 region and subsequent failures of replication have provoked considerable controversy. We here report six Welsh families multiply affected with schizophrenia in which there is no evidence for linkage between a dominant-like schizophrenia gene and 5q11-q13 markers. It is argued that our new results together with a combined reanalysis of previous studies suggest that a schizophrenia susceptibility gene can be excluded from the 5q11-q13 region. The apparent disparities between published results are most likely to reflect a chance finding in the one positive study and probably should not be interpreted as resulting from true linkage heterogeneity.

Eight days of treatment with two low doses of tetrahydroaminoacridine (THA), given once daily, substantially improved radial maze performance in two groups of rats which showed persistent deficits either after ibotenic acid lesions at the source of forebrain cholinergic projections, or after 28 weeks treatment with alcohol (20% v/v) in drinking water. However, in immature, aged or aged and alcohol-treated rats, acetylcholine content was not significantly affected in any of the brain areas measured, even though the treatment regime had proved behaviourally effective. Inhibition of brain acetylcholinesterase activity was only marginally increased by this treatment regime. Thus, if THA influences behaviour by enhancing cholinergic transmission, its effects do not appear to be related to its activity as a cholinesterase inhibitor, and alternative mechanisms of action should be investigated.

The choline-transport system has been solubilized from synaptic plasma membrane by using either sodium cholate or Triton X-100, and re-incorporated into unilamellar liposomes by using the technique of cholate dialysis. The criteria of choline-transport activity were saturability by excess choline, inhibition by hemicholinium-3, and trans-activation (i.e. stimulation of the uptake of [3H]choline into liposomes by preloading them with non-radioactive choline). Liposomes prepared from detergent extracts of synaptic plasma membrane and added lipid showed uptake of [3H]choline fulfilling these three criteria. Data on choline-transport activity of liposomes at various choline concentrations could be interpreted as implying that the transport system has two apparent Km values (2-5 microM and 50-100 microM), or alternatively that the system is composed of two or more negatively co-operating subunits (or units). It was shown by t.l.c. that the transported radioactivity was choline and that it was not significantly acetylated. Replacing Na+ by K+ on the outside of these liposomes partially inhibited uptake, and the formation of a potential gradient (inside negative) with valinomycin increased the total but not the saturable components of uptake when liposomes were prepared in a K+ medium, and transferred to an Na+ medium.