Elevation of Serum Dopamine-β-hydroxylase Activity with Forced Immobilization

THE formation of the neurotransmitter noradrenaline from 3,4-dihydroxyphenylethylamine (dopamine) is catalysed by the enzyme dopamine-β-hydroxylase (DBH)¹. This enzyme is associated both with the catecholamine-containing chromaffin granules in the adrenal medulla^2,3 and with the vesicular structures in sympathetic nerve terminals which contain catecholamines⁴. Furthermore, DBH activity is released with catecholamines into the perfusate after stimulation of either the isolated perfused adrenal gland⁵ or the isolated perfused spleen^6–8. DBH activity has been reported in the serum of both man and the rat^9,10. This activity is similar to adrenal and sympathetic nerve DBH activity with regard to cofactor requirements, oxygen requirement and kinetic characteristics^9,10. It has been suggested that serum DBH activity might be present as a result of release of enzyme with catecholamines from the adrenal glands and sympathetic nerves. If this is the case, serum DBH activity might be a useful and convenient index of sympathetic-adrenal activity. The work described here was undertaken to investigate both the source of the serum DBH and the effect on this activity of forced immobilization, a procedure which has been used as a model of stress and which has been shown to release catecholamines from the adrenal gland and increase catecholamine excretion¹¹.     

Isolation and Characterization of Coal Solubilizing Aerobic Microorganisms from Salt Range Coal Mines,Pakistan

Microbial solubilization of coal has been considered as a promising technology to convert raw coal into valuable products. In the present study, initially a total of 50 different aerobic bacterial and fungal isolates have been isolated from soil, coal and water samples of Dulmial Coal Mines, Chakwal, Pakistan, but on the basis of solubilization potential, only four isolates were selected for further study. The intensity of biosolubilization was measured by determining the weight loss of the coal pieces, which was observed to be about 25.93% by Pseudomonas sp. AY2, 36.36% by Bacillus sp. AY3 and 50% by Trichoderma sp. AY6, while Phanerochaete sp. AY5 showed maximum coal solubilization potential i.e. 66.67% in 30 days. UV/Vis spectrum revealed an increase in the pattern of absorbance of all treated samples compared to control referring to solubilization. Fourier transform infrared spectroscopy indicated alterations in the structure of treated coal in comparison to control coal suggesting breakdown in the complex structure of coal. The major absorbance bands in infrared spectroscopy for solubilization product were attributed to carbonyl (1,600 cm^?1), hydroxyl (3,450 cm^?1), cyclane (2,925 cm^?1), ether linkage (1,000–1,300 cm^?1), carboxyl (3,300–2,500 cm^?1) and side chains of aromatic ring (1,000–500 cm^?1). The presence of microorganisms and surface erosion of coal residues compared to control samples were observed by scanning electron microscopy, which suggested that isolated microorganisms were able to survive in coal for a longer period of time. Therefore, the present study concluded that microorganisms isolated from coal mines have excellent potential for coal solubilization which is considered as a crucial step in coal methanogenesis allowing them to be used successfully for in situ methane production to meet future energy demands.     

γ-Aminobutyric Acid Uptake by Sympathetic Ganglia

EXOGENOUS γ-aminobutyric acid (GABA) accumulates against a concentration gradient in isolated mammalian nervous tissue^1–3 and mixes with GABA stored in the tissue⁴. Thus, neurones which use GABA as an inhibitory transmitter might be identified by locating sites of accumulation of radioactively-labelled GABA using autoradiography^5–7, assuming that exogenous GABA is only taken up into neurones already containing GABA. A correlation between GABA uptake and endogenous content has been noted in slices from different parts of the brain³ and in different nerve-ending fractions^8–10. These experiments, however, do not show whether GABA can be accumulated in nerve tissue totally devoid of “gabanergic” neurones. To test this, we have measured the uptake of GABA by isolated sympathetic ganglia. The principal transmitter in the ganglion is acetylcholine while the postganglionic neurones are mainly adrenergic. By analogy with the brain, the ganglion contains negligible amounts of GABA, glutamic decarboxylase or GABA-transaminase^11,12.     

Cyclic AMP-dependent signaling system is a primary metabolic target for non-thermal effect of microwaves on heart muscle hydration

Previously, we have suggested that cell hydration is a universal and extra-sensitive sensor for the structural changes of cell aqua medium caused by the impact of weak chemical and physical factors. The aim of present work is to elucidate the nature of the metabolic messenger through which physiological solution (PS) treated by non-thermal (NT) microwaves (MW) could modulate heart muscle hydration of rats. For this purpose, the effects of NT MW–treated PS on heart muscle hydration, [³H]-ouabain binding with cell membrane, ⁴⁵Ca²⁺ uptake and intracellular cyclic nucleotides contents in vivo and in vitro experiments were studied. It is shown that intraperitoneal injections of both Sham-treated PS and NT MW–treated PS elevate heart muscle hydration. However, the effect of NT MW–treated PS on muscle hydration is more pronounced than the effect of Sham-treated PS. In vitro experiments NT MW–treated PS has dehydration effect on muscle, which is not changed by decreasing Na⁺ gradients on membrane. Intraperitoneal injection of Sham- and NT MW–treated PS containing ⁴⁵Ca²⁺ have similar dehydration effect on muscle, while NT MW–treated PS has activation effect on Na⁺/Ca²⁺ exchange in reverse mode. The intraperitoneal injection of NT MW–treated PS depresses [³H]-ouabain binding with its high-affinity membrane receptors, elevates intracellular cAMP and decreases cGMP contents. Based on the obtained data, it is suggested that cAMP-dependent signaling system serves as a primary metabolic target for NT MW effect on heart muscle hydration.     

The proteoglycans of bovine nasal cartilage and human articular cartilage: a sedimentation equilibrium analysis

Cartilage proteoglycan was isolated from bovine nasal septum and fractionated according to buoyant density after dissociative CsCl density gradient centrifugation. Gel-exclusion chromatography showed that hyaluronic acid was present in fractions of density lower than 1.69 g/mL. The molecular weight, assessed by sedimentation equilibrium analysis, of the proteoglycan present in the fractions with density > 1.69 g/mL, which appeared chromatographically homogeneous and constituted 54% of the preparation, ranged from 1.0 to 2.6 × 10⁶ for v = 0.55 cm³ g^?1. Carbodiimide-induced modification of the carboxyl groups by methylamine resulted in a reduction of the molecular weight to 0.74 – 1.25 × 10⁶. An analogous reduction in molecular weight was obtained after equilibration of this proteoglycan fraction with hyaluronic acid oligomers containing five disaccharide units. Since both procedures are known to cause inhibition of the interaction between proteoglycans and hyaluronic acid, it is suggested that this lower molecular-weight range represents the true degree of polydispersity of the sub-units of hyaline cartilage proteoglycan constituting this fraction, while the higher values obtained for the intact proteoglycan are the result of the presence of hyaluronic acid in the sample. The molecular-weight range of the whole proteoglycan subunit preparation, assessed after carboxyl group modification, was 0.5–1.2 × 10⁶. Apparently normal and abnormal cartilage was excised from single human osteoarthrosic femoral heads. Proteoglycans extracted by 4M guanidine hydrochloride were isolated after dissociative density gradient centrifugation and subjected to carboxyl group modification. Preparations from normal tissue exhibited molecular-weight averages ranging from 5 to 9 × 10⁵. A molecular-weight reduction was observed with proteoglycans isolated from abnormal areas.     

Polyanion-mediated mineralization — mineralization in coccolithophore (Pleurochrysis carterae) variants which do not express PS2, the most abundant and acidic mineral-associated polyanion in wild-type cells

Summary Highly acidic macromolecules are postulated intermediates in biomineralization because they sequester large numbers of calcium ions and occur in high concentrations at mineralizing foci in distantly related organisms. The mineral-associated polyanions in the coccolithophorePleurochrysis carterae are acidic polysaccharides which surround calcite elements on mineralized scales called coccoliths. PS2 is the most abundant and acidicPleurochrysis polyanion, containing four carboxyl groups per disaccharide repeat. This study examines the mineralizing function of PS2 by analysis of calcium carbonate deposition in PS2-deficient cells. Mineral-deficientPleurochrysis variants were separated from wild-type cells by density gradient centrifugation and analyzed for PS2 expression. Three independent low-density variants (two spontaneous and one chemically-induced) not expressing PS2 (ps2⁻) were isolated. The mineral content of each ps2⁻ variant was less than 5% of the wild-type level. Most ps2⁻ coccoliths were unmineralized, but when crystals were present, they occurred on the coccolith rim as in wild-type cells. When ps2⁻ rim elements were adequately large and numerous, they displayed the alternating A and B anvil shapes characteristic of wild-type rim elements. Although ps2⁻ cells may have defects which affect the expression of cellular components in addition to PS2, correlation of PS2 nonexpression with mineral deficiency in three independent variants supports the hypothesis that PS2 is a functional intermediate in mineral deposition. PS2 may affect the level ofPleurochrysis mineralization by (a) controlling the amount of calcium ions available for mineralization, (b) accelerating the rate of calcite nucleation, or (c) both. However, PS2 does not influence crystal morphology, since the more mature crystals in ps2⁻ cells exhibit the anvil-shaped morphology of wild-type crystals.     

Peripheral Neuropathy and Experimental Myeloma in the Mouse

SEVERAL forms of generalized and localized peripheral neuropathy have been associated in man with solitary or multiple myeloma^1–3. The cause of the nerve damage, which usually involves segmental demyelination^{4, 5} (that is, a specific lesion of the Schwann cell-myelin sheath) as well as some total nerve fibre degeneration, is unknown and so is its relationship to the protein produced by the abnormal plasma cells. The disorder has been classed as one of the “non-metastatic neurological complications of neoplasms”⁶, a group of mysterious disorders, almost all of unknown cause, which have not so far been reproduced in experimental animals. Recently, histological studies on the peripheral nerves of mice bearing transplanted plasma cell myelomas have shown that tumours secreting a particular type of protein are associated with a similar mixed peripheral neuropathy.     

Substance P antagonists and the role of tachykinins in non-cholinergic bronchoconstriction

Electrical field stimulation of guinea-pig isolated hilus bronchi induced tetrodotoxin-sensitive contractions of which only a minor part could be inhibited by atropine. The remaining non-cholinergic bronchonstriction was antagonized by a heptapeptide and an undecapeptide substance P (SP) analogue (Arg⁵, D-Trp^{7, 9}) SP_5–11, IC50 = 24.0 μM and (D-Pro², D-Trp^{7, 9})SP, IC50 = 10.0 μM. Of the exogenously added tachykinins, both eledoisin (8 times) and physalaemin (3 times) were more potent bronchoconstrictors than SP. Pretreatment with the SP-analogues shifted concentration-response curves to the tachykinins to the right, eledoisin being most readily antagonized. (Arg⁵, D-Trp^{7, 9})SP_5–11 also antagonized the neural response more readily than that of SP. In addition, in the frog isolated sciatic nerve preparation the two SP-analogues were found to possess potent lidocaine-like neurodepressant actions which further complicated the interpretation of the neural inhibitory effects of these compounds. It is concluded that if a tachykinin contributes to non-cholinergic bronchoconstriction, an eledoisin-like peptide is a more likely candidate than SP itself. Since SP-antagonists may have local anaesthetic properties their value as tools in neurophysiology seems limited. Inferentially, the non-cholinergic bronchoconstrictive neurotransmitter remains to be identified.     

Membrane Topology of Human Presenilin-1 in SK-N-SH Cells Determined by Fluorescence Correlation Spectroscopy and Fluorescent Energy Transfer

Presenilin-1 (PS1) protein acts as passive ER Ca²⁺ leak channels that facilitate passive Ca²⁺ leak across ER membrane. Mutations in the gene encoding PS1 protein cause neurodegeneration in the brains of patients with familial Alzheimer’s disease (FAD). FADPS1 mutations abrogate the function of ER Ca²⁺ leak channel activity in human neuroblastoma SK-N-SH cells in vitro (Das et al., J Neurochem 122(3):487–500, 2012) and in mouse embryonic fibroblasts. Consequently, genetic deletion or mutations of the PS1 gene cause calcium (Ca²⁺) signaling abnormalities leading to neurodegeneration in FAD patients. By analogy with other known ion channels it has been proposed that the functional PS1 channels in ER may be multimers of several PS1 subunits. To test this hypothesis, we conjugated the human PS1 protein with an NH₂-terminal YFP-tag and a COOH-terminal CFP-tag. As expected YFP–PS1, and PS1–CFP were found to be expressed on the plasma membranes by TIRF microscopy, and both these fusion proteins increased ER Ca²⁺ leak channel activity similar to PS1 (WT) in SK-N-SH cells, as determined by functional calcium imaging. PS1–CFP was either expressed alone or together with YFP–PS1 into SK-N-SH cell line and the interaction between YFP–PS1 and PS1–CFP was determined by Förster resonance energy transfer analysis. Our results suggest interaction between YFP–PS1 and PS1–CFP confirming the presence of a dimeric or multimeric form of PS1 in SK-N-SH cells. Lateral diffusion of PS1–CFP and YFP–PS1 in the plasma membrane of SK-N-SH cells was measured in the absence or in the presence of glycerol by fluorescence correlation spectroscopy to show that both COOH-terminal and NH₂-terminal of human PS1 are located on the cytoplasmic side of the plasma membrane. Therefore, we conclude that both COOH-terminal and NH₂-terminal of human PS1 may also be oriented on the cytosolic side of ER membrane.     

Picosecond fluorescence kinetics in spinach chloroplasts at room temperature. Effects of Mg2+

Single-photon timing with picosecond resolution is used to investigate the effect of Mg²⁺ on the room-temperature fluorescence decay kinetics in broken spinach chloroplasts. In agreement with an earlier paper (Haehnel, W., Nairn, J.A., Reisberg, P. and Sauer, K. (1982) Biochim. Biophys. Acta 680, 161–173), we find three components in the fluorescence decay both in the presence and in the absence of Mg²⁺. The behavior of these components is examined as a function of Mg²⁺ concentration at both the F₀ and the F_max fluorescence levels, and as a function of the excitation intensity for thylakoids from spinach chloroplasts isolated in the absence of added Mg²⁺. Analysis of the results indicates that the subsequent addition of Mg²⁺ has effects which occur at different levels of added cation. At low levels of Mg²⁺ (less than 0.75 mM), there appears to be a decrease in communication between Photosystem (PS) II and PS I, which amounts to a decrease in the spillover rate between PS II and PS I. At higher levels of Mg²⁺ (about 2 mM), there appears to be an increase in communication between PS II units and an increase in the effective absorption cross-section of PS II, probably both of these involving the chlorophyll $\text{a}\text{b}$ light-harvesting antenna.     

Autoradiographic Localization of <Superscript>3</Superscript>H-GABA in Rat Retina

γ-AMINOBUTYRIC acid (GABA) is present in all layers of vertebrate retinae^1–3: in the rabbit retina it seems to be most concentrated in the ganglion cell layers² while in the frog it is concentrated primarily in cell layers which are rich in amacrine cells¹. Recent autoradiographic studies of the distribution of ³H-GABA in rat brain slices after incubation in vitro suggest that the labelled amino-acid is selectively concentrated by certain neural elements^4,5. In a study of the distribution of ³H-GABA in rabbit retina after injection of the labelled amino-acid into the eye, Ehinger⁶ found that radioactivity was accumulated principally in the inner plexiform, inner nuclear and ganglion cell and nerve fibre layers. Labelling was also concentrated in some cells occupying the same position as amacrine cells and in some nerve cells of the ganglion cell layer.     

Structural and functional relationships in developing Pinus silvestris chloroplasts

The light dependent chloroplast development of dark grown seedlings of Pinus silvestris L. was followed by analyses of chlorophyll content, chlorophyll a/b ratios, chlorophyll/P700 ratios, chlorophyll-protein complexes and structural changes. Low-temperature fluorescence emission spectra of isolated chloroplasts and separation of sodium dodecyl sulphate solubilized chlorophyll-protein complexes by gel electrophoresis showed that the chlorophyll-protein complexes of photosystem 1 (P700-CPa), photosystem II (PS II-CPa) and the light-harvesting complex LH–CPa/b were present in dark grown seedlings. The low-temperature fuoorescence emission maxima of isolated P700–CPa and PS II–CPa shifted towards longer wavelengths during greening in light, indicating a light induced change of the chlorophyll organisation in the two photosystems. Illumination caused LH–CPa/b to increase relative to P700–CPa, whereas the ratio between LH–CPa/b and PS II–CPa remained essentially constant. Analyses of low-temperature fluorescence spectra with or without 0.01 M Mg²⁺ showed that the Mg²⁺ controlled distribution of excitation energy into PS I was activated upon illumination of the seedlings. The photosynthetic unit size, as defined by the chlorophyll/P700 ratio, did not change over a 96 h illumination period, although the chlorophyll content increased about 6–fold during that time. This result and the constant electron transport rate per unit chlorophyll and time during chlorophyll accumulation provided evidence for a sequential development of the photosynthetic units when illuminating dark grown pine cotyledons. Electron micrographs showed that exposure of dark grown seedlings to light for 2 h caused the prolamellar body to disappear and grana to form. These changes occurred prior to substantial accumulation of chlorophyll or change in the ratio between LH–CPa/b and P700–CPa. However, both the water-splitting system of photosystem II and the Mg²⁺ controlled redistribution of excitation energy was activated during this period.     

Cation-induced superoxide generation in tobacco cell suspension culture is dependent on ion valence

There have been many reports suggesting the involvement of reactive oxygen species (ROS), including superoxide anion (O₂^.–), in salt stress. Herein, direct evidence that treatments of cell suspension culture of tobacco (Nicotiana tabacum L.; cell line, BY‐2) with various salts of trivalent, divalent and monovalent metals stimulate the immediate production of O₂^.– is reported. Among the salts tested, LaCl₃ and GdCl₃ induced the greatest responses in O₂^.– production, whereas CaCl₂ and MgCl₂ showed only moderate effects; salts of monovalent metals such as KCl and NaCl induced much lower responses, indicating that there is a strong relationship between the valence of metals and the level of O₂^.– production. As the valence of the added metals increased from monovalent to divalent and trivalent, the concentrations required for maximal responses were lowered. Although O₂^.– production by NaCl and KCl required high concentrations associated with hyperosmolarity, the O₂^.– generation induced by NaCl and KCl was significantly greater than that induced simply by hyperosmolarity. Since an NADPH oxidase inhibitor, diphenyleneiodonium chloride, showed a strong inhibitory effect on the trivalent and divalent cation‐induced generation of O₂^.–, it is likely that cation treatments activate the O₂^.–‐generating activity of NADPH oxidase.     

Lack of Anti-macrophage Species Specificity in Anti-lymphocytic Serum

THE mode of action of anti-lymphocytic serum (ALS) in suppressing cellular and humoral immune responses remains uncertain^1,2 although there is evidence that it acts preferentially on a sub-population of thymus derived small lymphocytes^3–5. Its action, however, is not confined to lymphocytes but it also affects neutrophils⁶, epithelial cells⁷ and macrophages^8–14. On the other hand, specific anti-macrophagic activity of ALS has been either questioned¹⁵ or refuted^16–18. Anti-neutrophil activity of ALS may be responsible for the anti-inflammatory properties of ALS¹⁹ which have been shown to be separate from its immuno-suppressive activity and not to prolong graft survival²⁰. Macrophages and monocytes, however, play an important role in the immune response²¹ and anti-macro phage activity might account for much of the immuno-suppressive capacity of ALS. Previous reports indicated that ALS was species but not strain specific with respect to both anti-lymphocytic⁸ and anti-macrophagic activity¹³. Species specificity of anti-lymphocytic activity is, however, not absolute^22,33,34; and this report demonstrates that the same is true for anti-macrophage activity, particularly between related species.     

RNA Polymerase III of <Emphasis Type="Italic">Blastocladiella emersonii</Emphasis> is Mitochondrial

MULTIPLE RNA polymerases have been shown to exist in a wide variety of eukaryotic organisms^1–5. Two nuclear polymerases have been found in all the cells studied, each with a specific location and a specific function: the DEAE fraction I enzyme is located in the nucleolus and may be involved in the synthesis of ribosomal RNA^1,2,5,6; the DEAE fraction II enzyme is located in the non-nucleolar nucleoplasm and functions in the synthesis of DNA-like RNA^2–5,7. The DEAE fraction III enzyme was reported to exist in sea urchin¹, the aquatic fungus B. emersonii⁵ and to be present sometimes in rat liver preparations^1,8. Although there have been some reports that polymerase III is nuclear, Horgen and Griffin⁵ showed that the enzyme was sensitive to the prokaryotic RNA polymerase inhibitor rifampicin. They suggested that the fraction III enzyme may be mitochondrial, formed as the result of organelle contamination in their crude nuclear preparations. The results of this study show that the DEAE fraction III enzyme in B. emersonii is a mitochondrial enzyme, most likely functioning in the synthesis of mitochondrial RNA. The rifampicin sensitivity of the enzyme is further evidence of a prokaryotic origin of mitochondria^9,10.     

Insulin and Amino-acid Regulation of Polysomes in Perfused,Diabetic Rat Liver

IN normal mammalian liver, protein synthesis and concomitant polyribosome assembly are controlled by amino-acids^1–4. A similar control may be exerted by insulin in liver⁵ and muscle⁶. Both the effect of amino-acids on liver and that of insulin on muscle can be demonstrated with isolated, perfused tissue^4,7, but the interrelationship between these agents has not been clarified either in liver or in muscle. We have used perfused livers from diabetic rats to establish whether amino-acids and insulin promote polysomal assembly independently or in concert.     

Characterization of P700 as a photochemical quencher in isolated Photosystem I particles using simultaneous measurements of absorbance changes at 830 nm and photoacoustic signal

The relationship between the redox state of P700, the primary donor of PS I, monitored using absorbance changes at 830 nm and photochemical energy storage in PS I reaction centers assayed with the photoacoustic method (PA) was studied in isolated PS I submembrane particles aspirated onto nitrocellulose filters. Several donors have been used to support the electron transport through PS I. NADPH and NADH demonstrated low rates of electron donation to PS I, while ascorbate and ascorbate plus 2,6-dichlorophenolindophenol (DCIP) couple have been found more effective in both P700⁺ reduction and stimulation of the variable component of the PA signal. A linear relationship was found in isolated PS I particles between the (A_830,max – A_830,steady)/A_830,max and (PA_max – PA_steady)/PA_max ratios, which characterized the relative amount of P700 in the reduced state and the relative magnitude of the variable PA component, respectively. That linear relationship was obtained independently from the nature of electron donor used for the reduction of P700⁺. Such linear relationship was also obtained at various wavelengths of modulated light in the range of 660 to 720 nm, only the slope of the linear fits varied with wavelength. It is concluded that reduced P700 act as a photochemical quencher of absorbed energy. Variable thermal dissipation in PS I reaction centers of isolated submembrane particles linearly depends on the amount of reduced P700 and thus constitutes an appropriate indicator of the redox pressure applied to PS I. This revised version was published online in June 2006 with corrections to the Cover Date.     

Characterization of a resolved oxygen-evolving Photosystem II preparation from spinach thylakoids

An oxygen-evolving Photosystem (PS) II preparation was isolated after Triton X-100 treatment of spinach thylakoids in the presence of Mg²⁺. The structural and functional components of this preparation have been identified by SDS-polyacrylamide gel electrophoresis and sensitive spectrophotometric analysis. The main findings were: (1) The concentration of the primary acceptor Q of PS II was 1 per 230 chlorophyll molecules. (2) There are 6 to 7 plastoquinone molecules associated with a ‘quinone-pool’ reducible by Q. (3) The only cytochrome present in significant amounts (cytochrome b-559) occurred at a concentration of 1 per 125 chlorophyll molecules. (4) The only kind of photochemical reaction center complex present was identified by fluorescence induction kinetic analysis as PS II_α. (5) An E_m = ? 10 mV has been measured at pH 7.8 for the primary electron acceptor Q_α of PS II_α. (6) With conventional SDS-polyacrylamide gel electrophoresis, the preparation was resolved into 13 prominent polypeptide bands with relative molecular masses of 63, 55, 51, 48, 37, 33, 28, 27, 25, 22, 15, 13 and 10 kDa. The 28 kDa band was identified as the PS II light-harvesting chlorophyll $\text{a}\text{b-}\text{protein}$. In the presence of 2 M urea, however, SDS-polyacrylamide gel electrophoresis showed seven prominent polypeptides with molecular masses of 47, 39, 31, 29, 27, 26 and 13 kDa as well as several minor components. CP I under identical conditions had a molecular mass of 60–63 kDa.     

Morphine and Nalorphine impair Neuromuscular Transmission

THE narcotic analgesic morphine and the analgesic-antagonist nalorphine both depress the twitch and tetanus of the coaxially stimulated guinea-pig ileum¹. There is general agreement that morphine impairs the release of acetylcholine at this peripheral muscarinic site^1–4. Less is agreed about the effects of morphine at nicotinic sites^1,5–7. Because there is no evidence that the mechanisms of transmitter release differ fundamentally between muscarinic and nicotinic sites, we have examined the effects of morphine and nalorphine on some isolated skeletal neuromuscular preparations. A preliminary account of a portion of this work has already been reported⁸.     

Endogenous expression of FAD-linked PS1 impairs proliferation,neuronal differentiation and survival of adult hippocampal progenitors

Background

Alzheimer’s disease (AD) is characterized by progressive memory loss and impaired cognitive function. Early-onset familial forms of the disease (FAD) are caused by inheritance of mutant genes encoding presenilin 1 (PS1) variants. We have demonstrated that prion promoter (PrP)-driven expression of human FAD-linked PS1 variants in mice leads to impairments in environmental enrichment (EE)-induced adult hippocampal neural progenitor cell (AHNPC) proliferation and neuronal differentiation, and have provided evidence that accessory cells in the hippocampal niche expressing PS1 variants may modulate AHNPC phenotypes, in vivo. While of significant interest, these latter studies relied on transgenic mice that express human PS1 variant transgenes ubiquitously and at high levels, and the consequences of wild type or mutant PS1 expressed under physiologically relevant levels on EE-mediated AHNPC phenotypes has not yet been tested.

Results

To assess the impact of mutant PS1 on EE-induced AHNPC phenotypes when expressed under physiological levels, we exposed adult mice that constitutively express the PSEN1 M146V mutation driven by the endogenous PSEN1 promoter (PS1 M146V “knock-in” (KI) mice) to standard or EE-housed conditions. We show that in comparison to wild type PS1 mice, AHNPCs in mice carrying homozygous (PS1 ^M146V/M146V ) or heterozygous (PS1 ^M146V/+ ) M146V mutant alleles fail to exhibit EE-induced proliferation and commitment towards neurogenic lineages. More importantly, we report that the survival of newborn progenitors are diminished in PS1 M146V KI mice exposed to EE-conditions compared to respective EE wild type controls.

Conclusions

Our findings reveal that expression at physiological levels achieved by a single PS1 M146V allele is sufficient to impair EE-induced AHNPC proliferation, survival and neuronal differentiation, in vivo. These results and our finding that microglia expressing a single PS1 M146V allele impairs the proliferation of wild type AHNPCs in vitro argue that expression of mutant PS1 in the AHNPC niche impairs AHNPCs phenotypes in a dominant, non-cell autonomous manner.