Lactic acid bacteria with potential to eliminate fungal spoilage in foods.

AIMS: To investigate antifungal activity produced by lactic acid bacteria (LAB) isolated from malted cereals and to determine if such LAB have the capacity to prevent fungal growth in a particular food model system. METHODS AND RESULTS: The effect of pH, temperature and carbon source on production of antifungal activity by four LAB was determined. Pediococcus pentosaceus was used to conduct a trial to determine if it is feasible to eliminate Penicillium expansum, the mould responsible for apple rot, using an apple model. Penicillium expansum was incapable of growth during the trial on apple-based agar plates inoculated with the antifungal-producing culture, whereas the mould did grow on apple plates inoculated with an LAB possessing no antifungal activity. CONCLUSION: Partial characterization of the antifungal compounds indicates that their activity is likely to be because of production of antifungal peptides. The trial conducted showed that the antifungal culture has the ability to prevent growth of the mould involved in apple spoilage, using apples as a model. SIGNIFICANCE AND IMPACT OF THE STUDY: The ability of an LAB to prevent growth of Pen. expansum using the apple model suggests that these antifungal LAB have potential applications in the food industry to prevent fungal spoilage of food.

Utility of the MMPI-2 in assessing gender dysphoric patients.

We compared MMPI-2 (Butcher, Dahlstrom, Graham, Tellegen, & Kaemmer, 1989) profiles of 2 groups of adult biological men requesting sex reassignment surgery; 1 group was diagnosed with Diagnostic and Statistical Manual of Mental Disorders (3rd ed., rev. [DSM-III-R]; American Psychiatric Association, 1987) transsexualism and the other with gender identity disorder of adolescence and adulthood, nontranssexual type (GIDAANT). Although the mean profiles for the transsexual group did not demonstrate any psychopathology, the GIDAANT group showed moderate psychopathology. A cluster analysis indicated that 85% of the transsexual group showed low psychopathology and 47% of the GIDAANT group showed severe psychopathology. Neither the MMPI-2 results nor the DSM-III-R clinical evaluation support the conclusion of many authors that transsexualism is associated with severe personality disorder; rather, the data indicate that transsexualism and other gender identity disorders without persistent wish for sex reassignment differ significantly in degree of psychopathology.

Localization of M(2) muscarinic acetylcholine receptor protein in cholinergic and non-cholinergic terminals in rat hippocampus.

The muscarinic receptor family (M(1)-M(4)) mediates cholinergic modulation of hippocampal transmission. Pharmacological and physiological studies have indicated that a presynaptic receptor on cholinergic terminals plays a key role in regulating ACh release, although the molecular identity of this subtype is uncertain. In this study, the localization of the M(2) receptor is described in detail for the pyramidal cell layer in the CAl region of the hippocampus. Electron microscopic analysis of M(2) immunoreactivity in this area revealed mainly presynaptic expression of this subtype. Double-labeling experiments using antibodies to M(2) and to the vesicular acetylcholine transporter, a novel, specific marker of cholinergic terminals, were used to investigate the nature of these presynaptic receptors. These studies have revealed that M(2) is located in cholinergic and non-cholinergic terminals. This is the first direct anatomical evidence that suggests that M(2) may indeed function as a cholinergic autoreceptor in the hippocampus. The distribution of the M(2) receptor in non-cholinergic terminals also suggests functional roles for M(2) as a presynaptic heteroreceptor.

Activation of group II metabotropic glutamate receptors inhibits synaptic excitation of the substantia Nigra pars reticulata.

Loss of nigrostriatal dopaminergic neurons in Parkinson's disease (PD) leads to increased activity of glutamatergic neurons in the subthalamic nucleus (STN). Recent studies reveal that the resultant increase in STN-induced excitation of basal ganglia output nuclei is responsible for the disabling motor impairment characteristic of PD. On the basis of this, it is possible that any manipulation that reduces activity at excitatory STN synapses onto basal ganglia output nuclei could be useful in the treatment of PD. We now report that group II metabotropic glutamate receptors (mGluRs) are presynaptically localized on STN terminals and that activation of these receptors inhibits excitatory transmission at STN synapses. In agreement with the hypothesis that this could provide a therapeutic benefit in PD, a selective agonist of group II mGluRs induces a dramatic reversal of catalepsy in a rat model of PD. These results raise the exciting possibility that selective agonists of group II mGluRs could provide an entirely new approach to the treatment of PD. These novel therapeutic agents would provide a noninvasive pharmacological treatment that does not involve the manipulation of dopaminergic systems, thus avoiding the problems associated with current therapies.

Muscarinic-induced modulation of potassium conductances is unchanged in mouse hippocampal pyramidal cells that lack functional M1 receptors.

Activation of muscarinic acetylcholine (ACh) receptors (mAChRs) increases excitability of pyramidal cells by inhibiting several K+ conductances, including the after-hyperpolarization current (Iahp), the M-current (Im), and a leak K+ conductance (Ileak). Based on pharmacological evidence and the abundant localization of M1 receptors in pyramidal cells, it has been assumed that the M1 receptor is responsible for mediating these effects. However, given the poor selectivity of the pharmacological agents used to characterize these mAChR responses, rigorous characterization of the receptor subtypes that mediate these actions has not been possible. Surprisingly, patch clamp recording from CA1 pyramidal cells in M1 knockout mice revealed no significant difference in the degree of inhibition of Iahp, Im, or Ileak by the mAChR agonist, carbachol (CCh), as compared with wildtype controls. In addition, the M1-toxin was not able to block CCh's inhibition of the Iahp, Im, or Ileak These data demonstrate that the M1 receptor is not involved in increasing CA1 pyramidal cell excitability by mediating ACh effects on these K+ conductances.

Distribution and roles of metabotropic glutamate receptors in the basal ganglia motor circuit: implications for treatment of Parkinson's disease and related disorders.

The basal ganglia (BG) are a set of interconnected subcortical structures that play a critical role in motor control. The BG are thought to control movements by a delicate balance of transmission through two BG circuits that connect the input and output nuclei: the direct and the indirect pathways. The BG are also involved in a number of movement disorders. Most notably, the primary pathophysiological change that gives rise to the motor symptoms of Parkinson's Disease (PD) is the loss of dopaminergic neurons of the substantia nigra pars compacta (SNc) that are involved in modulating function of the striatum and other BG structures. This ultimately results in an increase in activity of the indirect pathway relative to the direct pathway and the hallmark PD symptoms of rigidity, bradykinesia, and akinesia. A great deal of effort has been dedicated to finding treatments for this disease. The current pharmacotherapies are aimed at replacing the missing dopamine, while the current surgical treatments are aimed at reducing transmission through the indirect pathway. Dopamine replacement therapy has proven to be helpful, but is associated with severe side effects that limit treatment and a loss of efficacy with progression of the disease. Recently developed surgical therapies have been highly effective, but are highly invasive, expensive, and assessable to a small minority of patients. For these reasons, new effort has been dedicated to finding pharmacological treatment options that will be effective in reducing transmission through the indirect pathway. Members of the metabotropic glutamate receptor (mGluR) family have emerged as interesting and promising targets for such a treatment. This review will explore the most recent advances in the understanding of mGluR localization and function in the BG motor circuit and the implications of those findings for the potential therapeutic role of mGluR-targeted compounds for PD.

Does long-term finasteride therapy affect the histologic features of benign prostatic tissue and prostate cancer on needle biopsy? PLESS Study Group. Proscar Long-Term Efficacy and Safety Study.

OBJECTIVES: Finasteride, a common agent used to treat benign prostatic hyperplasia (BPH), inhibits 5-alpha-reductase. Testosterone is converted by 5-alpha-reductase to the more potent dihydrotestosterone, which is the primary androgen in the prostate. Leuprolide is a stronger antiandrogen that is used to downstage prostate cancer before radical prostatectomy. Leuprolide induces marked atrophy of prostate carcinoma cells, which sometimes makes pathologic diagnosis of cancer difficult, although evaluation at radical prostatectomy is easier than at biopsy. It is unknown whether finasteride produces similar changes, which would result in greater diagnostic difficulty because such changes would be seen on biopsy to rule out cancer in men with suspicious clinical findings treated for BPH. The current study investigated the histologic effects of finasteride therapy on human prostate cancer and benign prostatic tissue on needle biopsy. METHODS: In blinded manner, we reviewed 53 needle biopsy specimens showing prostate carcinoma (35 treated with finasteride, 18 with placebo). Also reviewed in blinded manner were 50 benign needle biopsy specimens (25 treated with finasteride, 25 with placebo). The Gleason score, number of cores involved, percentage cancer involvement in a core, percentage of atrophic changes in cancer cells, presence of mitoses, blue-tinged mucinous secretions, prominent nucleoli, and high-grade prostatic intraepithelial neoplasia were documented for each case in the cancer group. The percentage of atrophy, basal cell hyperplasia, transitional metaplasia, chronic inflammation, and stromal proliferation was documented for each case in the benign group. RESULTS: No significant histologic differences were present in either the benign or cancer group between cases treated with finasteride and placebo. CONCLUSIONS: We conclude that finasteride treatment for BPH does not cause difficulty in the diagnosis of cancer in prostate needle specimens. It is possible that there are severely atrophic areas resulting from finasteride treatment that are undersampled. However, the conclusion that cancer seen on needle biopsy in men treated with finasteride is unaltered and readily identified as cancer remains valid.

Activation of NMDA receptors reverses desensitization of mGluR5 in native and recombinant systems.

The metabotropic glutamate receptor, mGluR5, has a critical role in induction of NMDA-receptor-dependent forms of synaptic plasticity and excitotoxicity. This is likely mediated by a reciprocal positive-feedback interaction between these two glutamate receptor subtypes in which activation of mGluR5 potentiates NMDA receptor currents and NMDA receptor activation potentiates mGluR5-mediated responses. We have investigated the mechanism by which NMDA receptor activation modulates mGluR5 function and find evidence that this response is mediated by activation of a protein phosphatase and a resultant dephosphorylation of protein kinase C phosphorylation sites on mGluR5. This form of neuromodulation may be important in a number of normal and pathological processes that involve activation of the NMDA receptor.

Muscarinic receptor subtypes involved in hippocampal circuits.

Muscarinic receptors modulate hippocampal activity in two main ways: inhibition of synaptic activity and enhancement of excitability of hippocampal cells. Due to the lack of pharmacological tools, it has not been possible to identify the individual receptor subtypes that mediate the specific physiological actions that underlie these forms of modulation. Light and electron microscopic immunocytochemistry using subtype-specific antibodies was combined with lesioning techniques to examine the pre- and postsynaptic location of m1-m4 mAChR at identified hippocampus synapses. The results revealed striking differences among the subtypes, and suggested different ways that the receptors modulate excitatory and inhibitory transmission in distinct circuits. Complementary physiological studies using m1-toxin investigated the modulatory effects of this subtype on excitatory transmission in more detail. The implications of these data for understanding the functional roles of these subtypes are discussed.

Active and passive immunization against HIV type 1 infection in chimpanzees.

Several vaccine strategies have been developed to prevent primary infection with human immunodeficiency virus (HIV), and some of the candidate vaccines have been tested in chimpanzees to determine their safety, efficacy, and to delineate immune correlates of protection. To date, more than 25 vaccines representing active and passive immunization strategies have been evaluated in the chimpanzee model. Efficacy of a given vaccine was based on protection against primary infection with HIV after intravenous or mucosal challenge with cell-free or cell-associated virus. Based on the results from a majority of the studies, neutralizing antibodies appear to play a major role in preventing primary infection with HIV.

Activation of the genetically defined m1 muscarinic receptor potentiates N-methyl-D-aspartate (NMDA) receptor currents in hippocampal pyramidal cells.

Evidence suggests that cholinergic input to the hippocampus plays an important role in learning and memory and that degeneration of cholinergic terminals in the hippocampus may contribute to the memory loss associated with Alzheimer's disease. One of the more prominent effects of cholinergic agonists on hippocampal physiology is the potentiation of N-methyl-D-aspartate (NMDA)-receptor currents by muscarinic agonists. Here, we employ traditional pharmacological reagents as well as m1-toxin, an m1 antagonist with unprecedented selectivity, to demonstrate that this potentiation of NMDA-receptor currents in hippocampal CA1 pyramidal cells is mediated by the genetically defined m1 muscarinic receptor. Furthermore, we demonstrate the colocalization of the m1 muscarinic receptor and the NR1a NMDA receptor subunit at the electron microscopic level, indicating a spatial relationship that would allow for physiological interactions between these two receptors. This work demonstrates that the m1-muscarinic receptor gene product modulates excitatory synaptic transmission, and it has important implications in the study of learning and memory as well as the design of drugs to treat neurodegenerative diseases such as Alzheimer's.

Differential presynaptic and postsynaptic expression of m1-m4 muscarinic acetylcholine receptors at the perforant pathway/granule cell synapse.

A family of muscarinic acetylcholine receptor proteins mediates diverse pre- and postsynaptic functions in the hippocampus. However the roles of individual receptors are not understood. The present study identified the pre- and postsynaptic muscarinic acetylcholine receptors at the perforant pathway synapses in rat brain using a combination of lesioning, immunocytochemistry and electron microscopic techniques. Entorhinal cortex lesions resulted in lamina-specific reductions of m2, m3, and m4 immunoreactivity in parallel with the degeneration of the medial and lateral perforant pathway terminals in the middle and outer thirds of the molecular layer, respectively. In contrast, granule cell lesions selectively reduced m1 and m3 receptors consistent with degeneration of postsynaptic dendrites. Direct visualization of m1-m4 by electron microscopic immunocytochemistry confirmed their differential pre- and postsynaptic localizations. Together, these findings provide strong evidence for both redundancy and spatial selectivity of presynaptic (m2, m3 and m4) and postsynaptic (m1 and m3) muscarinic acetylcholine receptors at the perforant pathway synapse.

Activation of latent transgenes in Arabidopsis using a hybrid transcription factor.

A hybrid transcription factor comprising a fusion of the DNA-binding domain of Saccharomyces cerevisiae GAL4 and the transcription activation domain of maize C1 was expressed in stably transformed Arabidopsis. Additional transgenic lines were created containing test genes controlled by a synthetic promoter consisting of concatemeric copies of the cis-acting site recognized by GAL4 (UASG) fused to a minimal promoter. The GAL4/C1 effector line was crossed to two lines containing a synthetic promoter/GUS fusion. Both histochemical staining and GUS activity assays indicate strong activation of GUS expression was achieved only after crossing. The GAL4/C1 effector line was also crossed to 15 lines containing a synthetic promoter/antisense adenylosuccinate synthetase gene. Severely retarded growth, and in some cases lethality, was observed in 40% of the F1 lines. This system of activation by crossing is generally useful for activating expression of test transgenes.

Generation of carbohydrate-deficient transferrin by enzymatic deglycosylation of human transferrin.

Carbohydrate-deficient transferrin (CDT) molecules are transferrin isoforms that lack one or both of the carbohydrate groups attached to a normal human transferrin molecule. CDT has been reported to be a sensitive and specific marker for diagnosing alcoholism. This report demonstrates the in vitro generation of CDT molecules that can potentially be used as the standard in measuring CDT concentrations. This was achieved by deglycosylation of human transferrin with the enzyme Endo-beta-N-acetylglucosaminidase F2 (Endo-F2). The enzyme was immobilized on sepharose beads, which were packed into a column. The immobilization of the enzyme not only eliminated the Endo-F2 contamination of CDT, but also rendered the enzyme suitable for repetitive use. In this manner, it was possible to obtain at least 200 mg of CDT over a period of more than 3 mo, without any noticeable decrease of enzyme activity, using only 3.0 micrograms of enzyme. This proved to be an efficient method for generating CDT.

Superinfection with human immunodeficiency virus type 2 can reactivate virus production in baboons but is contained by a CD8 T cell antiviral response.

An animal model was used to assess whether resistance to superinfection by human immunodeficiency virus (HIV) can exist in vivo. Asymptomatic baboons (Papio cynocephalus), previously infected with HIV-2, were first challenged with homologous virus (HIV-2UC2 or HIV-2UC14) and later with heterologous virus (HIV-2UC12). After both virus inoculations, either resistance to viral infection or a transient viremia was observed. The original virus was recovered in 3 baboons, suggesting that reactivation of a latent infection occurred on heterologous challenge and that HIV-2 superinfection is blocked by processes established during prior infection. Antibody titers measured by ELISA and virus neutralization remained at low levels. However, suppression of HIV-1 replication was observed with CD8 T cells and filtered cell culture supernatants. The soluble factor involved was not a beta-chemokine. This resistance to HIV superinfection appears to be mediated at least in part by CD8 T cells that suppress virus production.

Muscarinic acetylcholine receptor immunoreactivity after hippocampal commissural/associational pathway lesions: evidence for multiple presynaptic receptor subtypes.

The present study addressed the hypothesis that differential localization of ml-m4 subtypes in the inner one-third of the dentate molecular layer is due to selective presynaptic expression of the receptor proteins on the hippocampal commissural/associational pathways. Physical and chemical lesions of the commissural and associational pathways were used to denervate afferent terminals in the inner one-third of the molecular layer, and fluid injections were used to lesion granule cells, their postsynaptic target. Immunocytochemistry utilizing muscarine acetylcholine receptor (mAChR) subtype-specific antibodies was used to identify changes in expression patterns in the molecular layer postlesion. m1 immunoreactivity in the molecular layer did not change after commissural/associational pathway lesions. m2 immunoreactivity in the inner one-third of the molecular layer was attenuated only after lesions involving the associational pathway. In contrast, m3 and m4 immunoreactivity in the inner one-third of the molecular layer was almost completely abolished by lesions of both pathways simultaneously. Granule cell lesions greatly attenuated m1 and m3 immunoreactivity in the molecular layer, with little to no diminution of m2 and m4 immunoreactivity. The results indicate that, in the inner one-third of the molecular layer, m1 and m3 are mainly postsynaptic on granule cells, whereas m2 and m4 are presynaptic on the commissural/associational pathways. This study provides direct anatomical evidence for the diversity of molecular subtypes presynaptically on the commissural/associational pathways.

Reducing MMPI-2 defensiveness: the effect of specialized instructions on retest validity in a job applicant sample.

The MMPI-2 is often used for screening job applicants when public safety or security are at risk. Inherent in such applications is concern for profile validity and test defensiveness. In this study, we examine the impact of revised instructions on profile validity for a group of job applicants who initially produced invalid profiles. Participants were 271 male applicants for airline pilot positions. Of these, 72 produced invalid defensive MMPI-2 profiles during preemployment screening. The MMPI-2 was readministered to these applicants with instructions informing them of validity scales and instructing them to respond in a more open, honest manner. Comparisons were made between valid and invalid profiles for initial administrations and between valid and invalid profiles at readministration. Some clinical scales were more elevated for valid, nondefensive profiles. Most content scales showed more elevation for valid profiles, and 12% of the applicants who were retested produced significant elevations (T>or=65) on the content scales. Profiles were similar to those produced by employed pilots of a previous study.

Muscarinic acetylcholine receptor subtype, m2: diverse functional implications of differential synaptic localization.

The muscarinic acetylcholine receptor (mAChR) molecular subtype, m2, has been postulated to be the presynaptic cholinergic autoreceptor in many brain regions. However, due to a lack of subtype-specific pharmacological agents, conclusive evidence for m2 as an autoreceptor remains elusive. The development of subtype-specific antibodies has enabled extensive characterization of the synaptic localization of the m2 subtype. Specifically, double-labeling immunocytochemistry with m2 antibodies and antibodies to the vesicular acetylcholine transporter (VAChT), a novel specific marker of cholinergic terminals, in the striatum has allowed the first direct anatomical evidence of m2 localization in cholinergic terminals. Additionally, other anatomical studies in striatum and the septohippocampal pathway have revealed that this subtype is also expressed presynaptically in non-cholinergic terminals, and is postsynaptically expressed in both cholinergic and non-cholinergic neurons. The implications of these data for understanding the functional roles of this subtype are discussed.

Expression of m1-m4 muscarinic acetylcholine receptor immunoreactivity in septohippocampal neurons and other identified hippocampal afferents.

Muscarinic cholinergic transmission plays an important role in modulating hippocampal activity and many higher brain functions. Many of the modulatory effects of acetylcholine on hippocampal function result from direct effects in the hippocampus or from actions on the hippocampal afferent neurons. At each site, the differential expression of a family of five distinct but related receptor subtypes governs the nature of the response. The aim of the present study was to identify the subtypes expressed in the hippocampal afferent neurons by combining retrograde tracing with immunocytochemistry. The retrograde tracer, wheat germ agglutinin conjugated to horseradish peroxidase, was injected into the hippocampus unilaterally to label afferent neurons, and was combined with muscarinic (m) acetylcholine (ACh) receptors (mAChRs) with immunocytochemistry to identify the m1-m4 subtypes expressed. The retrogradely labeled cells in the basal forebrain that contribute to the septohippocampal pathway were found to express m2, m3, and, to a lesser extent, m1. Commissural/associational pathway neurons, which were identified by retrogradely labeled cells in the ipsi- and contralateral dentate gyrus, expressed m1, m3, and m4. The retrogradely labeled cells in the entorhinal cortex of the perforant pathway expressed predominantly m1 and m3, with fewer neurons expressing m2 and m4. Raphe-hippocampal cells were found to express m1. Thus, this study provides evidence for the diversity of mAChR subtypes expressed in neurons that project to the hippocampus. The complex modulation by acetylcholine of hippocampal function, therefore, is governed not only by the variety of mAChRs expressed in the hippocampus but also by their differential expression in extrinsic hippocampal afferents.