RNA- and ATAC-sequencing Reveals a Unique CD83+ Microglial Population Focally Depleted in Parkinson's Disease.

All brain areas affected in Parkinson's disease (PD) show an abundance of microglia with an activated morphology together with increased expression of pro-inflammatory cytokines, suggesting that neuroinflammation may contribute to the neurodegenerative process in this common and incurable disorder. We applied a single nucleus RNA- and ATAC-sequencing approach using the 10x Genomics Chromium platform to postmortem PD samples to investigate microglial heterogeneity in PD. We created a multiomic dataset using substantia nigra (SN) tissues from 19 PD donors and 14 non-PD controls (NPCs), as well as three other brain regions from the PD donors which are differentially affected in this disease: the ventral tegmental area (VTA), substantia inominata (SI), and hypothalamus (HypoTs). We identified thirteen microglial subpopulations within these tissues as well as a perivascular macrophage and a monocyte population, of which we characterized the transcriptional and chromatin repertoires. Using this data, we investigated whether these microglial subpopulations have any association with PD and whether they have regional specificity. We uncovered several changes in microglial subpopulations in PD, which appear to parallel the magnitude of neurodegeneration across these four selected brain regions. Specifically, we identified that inflammatory microglia in PD are more prevalent in the SN and differentially express PD-associated markers. Our analysis revealed the depletion of a CD83 and HIF1A- expressing microglial subpopulation, specifically in the SN in PD, that has a unique chromatin signature compared to other microglial subpopulations. Interestingly, this microglial subpopulation has regional specificity to the brainstem in non-disease tissues. Furthermore, it is highly enriched for transcripts of proteins involved in antigen presentation and heat-shock proteins, and its depletion in the PD SN may have implications for neuronal vulnerability in disease.

Extracellular Tau Oligomers Produce An Immediate Impairment of LTP and Memory.

Non-fibrillar soluble oligomeric forms of amyloid-ß peptide (oAß) and tau proteins are likely to play a major role in Alzheimer's disease (AD). The prevailing hypothesis on the disease etiopathogenesis is that oAß initiates tau pathology that slowly spreads throughout the medial temporal cortex and neocortices independently of Aß, eventually leading to memory loss. Here we show that a brief exposure to extracellular recombinant human tau oligomers (oTau), but not monomers, produces an impairment of long-term potentiation (LTP) and memory, independent of the presence of high oAß levels. The impairment is immediate as it raises as soon as 20 min after exposure to the oligomers. These effects are reproduced either by oTau extracted from AD human specimens, or naturally produced in mice overexpressing human tau. Finally, we found that oTau could also act in combination with oAß to produce these effects, as sub-toxic doses of the two peptides combined lead to LTP and memory impairment. These findings provide a novel view of the effects of tau and Aß on memory loss, offering new therapeutic opportunities in the therapy of AD and other neurodegenerative diseases associated with Aß and tau pathology.

Gene structure and promoter function of murine Munc18-2, a nonneuronal exocytic Sec1 homolog.

Sec1 family proteins are regulators of diverse exocytic processes, from yeast to man. Three mammalian homologues, Munc18-1, -2, and -3 have been described. We have studied the structure and expression of the mouse Munc18-2 gene. The Munc18-2 gene comprises 19 exons whose sizes range from 50 to 158 bp, with a total gene size of approximately 11 kb. A single transcript of 2.1 kb is expressed in multiple non-neuronal murine tissues. Munc18-2 has a striking resemblance to Munc18-1 in structure despite only 60% sequence identity, suggesting a recent gene duplication event. Analysis of the region upstream of the transcription start site shows that Munc18-2 has a TATA-less promoter, with a consensus initiator (Inr) sequence at the start of transcription, several Sp1 binding sites, and strong promoter activity in RBL-2H3 mast cells. The region from +5 to -430 is more active than +5 to -800, suggesting upstream repressor elements.

Growth rate related concentration changes of the starvation response regulators sigmaS and ppGpp in glucose-limited fed-batch and continuous cultures of Escherichia coli.

The entry of Escherichia coli into glucose starvation is connected to the induction of both the stringent response and the general stress response. The concentration of the corresponding response regulators ppGpp and sigmaS (RpoS, KatF) was investigated during glucose-limited growth at different growth rates and with different cultivation strategies, including fed-batch fermentation, chemostat, and a continuous-cultivation technique with computer-controlled decrease of the feeding rate (A-stat). This study confirms the model of the in vivo effect of ppGpp on sigmaS by the direct measurement of ppGpp and sigmaS levels. The response of both regulators was clearly shown in situations where a nutrient shift occurs. A correlation in the concentration of both regulators was also shown under steady-state conditions. However, during transients with slow continuous changes of the nutrient availability, this concerted reaction of ppGpp and sigmaS is less apparent, indicating the specific importance of these regulators for the adaptation of the cells to fast changes of environmental parameters.

Amplification of ColE1 related plasmids in recombinant cultures of Escherichia coli after IPTG induction.

ColE1-derived plasmids containing different recombinant genes which are controlled by the tac promoter were amplified following induction with IPTG, but no amplification occurred if product formation was not induced. The plasmid copy number of recombinant E. coli increased three- to sixfold within a period of about 6 h in shake flask experiments, batch cultures, and glucose-limited fed-batch cultivations. Plasmid amplification occurred in E. coli B strains as well as in K-12 strains with different plasmids (rop+ and rop-) coding for various heterologous proteins. The amplification was not caused by a toxic effect of IPTG, but was related to a strong inhibition of translation and chromosomal replication after the induction of heterologous gene expression. Similar to the amplification after chloramphenicol addition, plasmid replication proceeded even if oriC replication and translation were inhibited following strong induction of a recombinant gene. In accordance with the effect of chloramphenicol, the level of ppGpp, which is a negative regulator of ColE1 derived plasmid replication, decreased after induction.

The general stress sigma factor sigmaS of Escherichia coli is induced during diauxic shift from glucose to lactose.

The general stress sigma factor sigmaS (RpoS) of Escherichia coli is strongly induced in response to glucose starvation. This increase in the cellular sigmaS level is due to stabilization of sigmaS, which under non-stress conditions is subject to rapid proteolysis. In the present study, it is demonstrated that sigmaS is also induced during the diauxic shift from glucose to lactose, i.e., under conditions of glucose exhaustion in the presence of another, less-preferred carbon source that eventually gets utilized. This sigmaS induction, which is due to stabilization, is transient and precedes the induction of beta-galactosidase. In parallel, sigmaS-dependent genes are transiently activated, as was shown here for osmY. Although sigmaS can mediate transcription of lacZ in vitro, sigmaS does not contribute to the induction of beta-galactosidase during the diauxic lag phase. Rather, the induction of sigmaS and the general stress response during the diauxic shift plays the role of a rapidly activated emergency system, which is shut off again as soon as the cells are able to cope with the stress situation by utilizing a more specific and more economical system.

Ibotenic acid lesions in the amygdaloid central nucleus but not in the lateral subthalamic area prevent the acquisition of differential Pavlovian conditioning of bradycardia in rabbits.

The present study examined the effect of ibotenic acid lesions in the amygdaloid central nucleus (ACe) or in the lateral zona incerta of the subthalamus (LZI) on the acquisition of differential Pavlovian conditioning of bradycardia in rabbits. Previous work has shown that bilateral electrolytic lesions in either ACe or LZI abolished the retention of conditioned heart rate (HR) responses. In order to determine whether these findings were due to destruction of cells intrinsic to ACe or LZI, ibotenic acid lesions were placed bilaterally in either structure or in control sites. Following recovery, animals were subjected to differential Pavlovian conditioning in which one tone (CS+) was paired with periorbital shock and a second tone (CS-) was presented alone. It was found that destruction of cell bodies in ACe, but not LZI, prevented the acquisition of the differential bradycardiac conditioned response. In addition, ACe lesions did not interfere with baseline HR, the HR orienting response, the HR unconditioned response to shock, or the concomitantly conditioned corneoretinal potential. The results of this study suggest that destruction of cells intrinsic to ACe selectively prevents the acquisition of differentially conditioned HR, and perhaps other conditioned responses related to conditioned arousal, but does not affect unlearned HR responses or specific somatomotor conditioned responses.

Auditory cortex lesions prevent the extinction of Pavlovian differential heart rate conditioning to tonal stimuli in rabbits.

The present study examined the role of the auditory cortex in the extinction of differentially conditioned heart rate (HR) responses in rabbits. Lesions were placed bilaterally in either the auditory cortex or the visual cortex. Three days after recovery from surgery, the auditory cortex lesion group and the visual cortex lesion control group were habituated to the tonal conditioned stimuli (CSs), and then given 2 days of Pavlovian differential conditioning (60 trials per day) in which one tone (CS+) was always paired with the unconditioned stimulus and another tone (CS-) was never paired with the unconditioned stimulus. Animals that had demonstrated reliable differential conditioning (CS+ response at least 5 beats greater than the CS- response) were placed on an extinction schedule for 7 days. The extinction schedule was identical to the differential conditioning schedule with the exception that shock never followed the CS+. The results of the study indicate that auditory cortex lesions prevent the extinction of differential bradycardia conditioned responses (CRs) to tonal CSs. Whereas the bradycardia responses to the CS+ quickly extinguished in the group that had control lesions in the visual cortex, the auditory cortex lesion group continued to exhibit significantly larger bradycardiac HR CRs to the CS+ relative to the CS- during all 7 days of extinction. These results suggest that the animals in the auditory cortex lesioned group did not inhibit responses to a previously reinforced stimulus (i.e., CS+) as well as animals with control lesions in the visual cortex.

Role of auditory cortex in the acquisition of differential heart rate conditioning.

Previous findings from our laboratory indicate that lesions of the auditory cortex disrupt the retention of differentially conditioned bradycardiac responses to tonal stimuli in rabbits. In the present experiment, the effect of lesions of the auditory cortex on the acquisition of differential bradycardiac conditioning was examined. The effect of lesions in the auditory cortex were compared to the effect produced by control lesions in the visual cortex. After 7 days of recovery, animals received 7 days of differential Pavlovian bradycardiac conditioning in which one tone (CS+) was paired with the unconditioned stimulus, and another tone (CS-) was never paired with the unconditioned stimulus. All animals demonstrated differential conditioning during the first 3 days of conditioning. On days 4-7, however, auditory cortex lesioned animals did not exhibit significant differential heart rate (HR) conditioning, whereas control animals with lesions in the visual cortex showed no loss of conditioning during this period. The loss of differential conditioning in animals with lesions in the auditory cortex appears to be due to an increase in the magnitude of the response to the CS-. These data support the hypothesis that the auditory cortex serves to inhibit the response to the CS- in differential conditioning of bradycardia to acoustic stimuli, and that the inhibition may be mediated by a descending corticothalamic or corticolimbic pathway.

[Thermologic studies in inflammatory rheumatic diseases--general presentation and personal results in rheumatoid arthritis]. / Thermologische Untersuchungen bei entzündlich-rheumatischen Erkrankungen--allgemeine Darstellung und eigene Ergebnisse bei Rheumatoid-Arthritis.

The registration of the temperature of the body surface gains an increasing significance among others for the diagnosis of inflammatory rheumatic diseases. A criterion of the inflammation of the joint, the hyperthermy as well as the disturbances of the microcirculation may be made evident by means of the thermological methods. Own results in healthy persons and patients with rheumatoid arthritis as well as evidence of the literature confirm the possibilities of the measurements of temperature for the rheumatology. On account of the great inter- and intra-individual variability of the skin temperature an orientation at the temperatures of normal values of certain measuring regions is not reasonable. When the results of the temperature measurement are estimated, therefore, a comparison with temperature patterns of healthy persons must be done. Using dynamic measuring methods the evidence of thermological investigations may essentially be enlarged. The evaluation of the results of the measurement is increasingly performed with the help of computers.

The role of amygdaloid central nucleus in the retention of differential pavlovian conditioning of bradycardia in rabbits.

The present study examined the role of the amygdaloid central nucleus (ACE) in the retention of differential pavlovian conditioning of bradycardia in rabbits. Electrodes were implanted bilaterally in ACE or in control sites just dorsal and rostral to ACE. Following recovery, animals were subjected to differential pavlovian conditioning in which one tone (CS+) was paired with periorbital shock and a second tone (CS-) was presented alone. Subsequent electrolytic lesions abolished the heart rate (HR) conditioned response (CR), yet had no effect on HR orienting response, unconditioned response, or baseline. In a follow-up experiment, corneoretinal potential (CRP) and HR were recorded. Bilateral ACE lesions profoundly attenuated or abolished the HR CR without abolishing CRP CRs. The major finding of this study is that bilateral lesions of ACE selectively attenuate the HR CR while not necessarily abolishing other CRs.

Lateral subthalamic area as mediator of classically conditioned bradycardia in rabbits.

This study was conducted to examine whether a central efferent pathway passing through the lateral zona incerta (LZI) of the subthalamus selectively mediates the bradycardia conditioned response (CR) in rabbits. Electrodes were implanted bilaterally in LZI or in control sites just dorsal or ventral to LZI. Two days following surgery, animals were subjected to Pavlovian conditioning or to pseudoconditioning. Subsequent bilateral electrolytic lesions did not influence the heart rate (HR) orienting response, unconditioned response, baseline, or lack of response to pseudoconditioning. Bilateral LZI lesions alone abolished the HR CR. In a follow-up experiment, the corneoretinal potential (CRP) CR and HR were recorded. Bilateral LZI lesions following conditioning to a criterion of 65% CRP CRs abolished the HR CR without affecting CRP CRs. The present findings indicate that LZI is part of an efferent pathway that selectively mediates the HR CR in rabbits.

Effect of Cultural Practices, Soil Phosphorus, Potassium, and pH on the Incidence of Fusarium Head Blight and Deoxynivalenol Levels in Wheat.

In a survey conducted in 1984 in Essex, Lambton, and Middlesex Counties of Ontario, Canada, greater incidences of head blight and greater concentrations of deoxynivalenol in grain were observed in fields of winter wheat planted after corn than in those planted after soybeans, barley, and mixed grains. Neither head blight nor deoxynivalenol level were correlated significantly with soil P, K, or pH. Head blight was reduced in wheat planted after corn where the residues from a preceding corn crop were plowed down or where seed was treated with Vitaflo 280. However, head blight and deoxynivalenol levels were not affected significantly by the level of N application, cultivar, other diseases or herbicides.

Decline in deoxynivalenol (vomitoxin) concentrations in 1983 ontario winter wheat before harvest.

Samples of winter wheat were collected in 1983 from five fields in southwestern Ontario that were naturally infected with head blight. The geometric mean concentration of deoxynivalenol (vomitoxin) in the grain declined significantly from 1.56 mug/g on 7 July to 0.21 mug/g on 14 July and 0.11 mug/g on 18 July and did not increase at harvest (20 through 27 July).

Heritability of grain yield, plant height and test weight of a population of winter wheat adapted to Southwestern Ontario.

Grain yield, plant height and test weight were studied in a population of winter wheat (Triticum aestivum L.). The population consisted of F2 bulk populations of 42 crosses among 11 genotypes adapted to S. W. Ontario. Heritabilities were: 0.30±0.32 for yield, 0.77±0.15 for height and 0.98±0.08 for test weight. Predicted genetic gain with 10% selection intensity was 0.15 t/ha for yield, 10.1 cm for height and 3.00 kg/hl for test weight. The low heritability for yield indicates that effective selection would require pedigree information and progeny tests, while the high heritabilities for height and test weight indicate that selection for these traits using single plots would be appropriate.

Differentiation between nucellar and zygotic citrus seedlings by leaf shape.

Leaf length divided by leaf width, (L/W), was studied as a criterion for differentiating between nucellar and zygotic citrus seedlings. Progeny of crosses of the variety 'King' as a seedparent (known to produce both nucellar and zygotic seedlings) fell into two classes: one with L/W similar to 'King' and the other midway between 'King' and the pollen parent. In several zygotic seedling populations, population means were intermediate between those of the parents. L/W can be an effective criterion for differentiating between nucellar and zygotic citrus seedlings where the parents differ appreciably.