An exploration of the impact of SARS-CoV-2 (COVID-19) restrictions on marginalised groups in the UK.

BACKGROUND: To contain the spread of COVID-19 within the UK over the past year, there have been a series of local and national lockdowns. These restrictions are likely to have impacted upon the health and well-being of marginalised groups who rely on now closed social and community support services to stay healthy. An understanding of the experiences of marginalised people is important; therefore, this study aimed to explore the impact of the COVID-19 restrictions on the health and well-being of marginalised groups in the UK. METHODS: In summer 2020, a rapid telephone survey was conducted by trained, trusted volunteers with 76 participants who were from marginalised groups. As part of this survey, 64 participants consented to describe their experience of lockdown. These case studies were thematically analysed to identify patterns of meaning. RESULTS: Findings indicate that lockdown led to the deterioration of health of participants, impacted adversely on their socio-economic positions and affected access to food and essential supplies. In addition, government public health messaging was considered confusing and inadequate. CONCLUSIONS: This study highlights the need for pathways into services which support marginalised groups to remain accessible during periods of restrictions and essential supplies and food to be mapped and protected for marginalised individuals within our local communities.

A qualitative exploration of patients' experiences with and perceptions of recommencing feeding after colorectal surgery.

BACKGROUND: Many patients who undergo lower gastrointestinal surgery neither recommence feeding within timeframes outlined by evidence-based guidelines, nor meet their nutrition requirements in hospital. Given that the success of timely and adequate post-operative feeding is largely reliant on patient adherence, the present study explored patients' perceptions of recommencing feeding after colorectal surgery to determine areas of improvement to meet their needs and expectations. METHODS: This qualitative study involved one-on-one, semi-structured interviews with patients receiving care after colorectal surgery in an Australian tertiary teaching hospital. Purposive sampling was used to ensure maximal variation in age, sex, procedural type and post-operative nutrition care experience. Interviews were audio recorded, with data transcribed verbatim before being thematically analysed. Emergent themes and subthemes were discussed by all investigators to ensure consensus of interpretation. RESULTS: Sixteen patients were interviewed (female 56%; age 61.5 ± 12.3 years). Three overarching themes emerged from the data: (i) patients make food-related decisions based on ideologies, experience and trust; (ii) patients appreciate the opportunity to participate in their nutrition care; and (iii) how dietary information is communicated influences patients' perceptions of and behaviours towards nutrition. CONCLUSIONS: Enabling patients to select from a wide range of foods from post-operative day 1 (by prescribing an unrestricted diet in line with evidence-based practice guidelines) in conjunction with delivering clear, simple and encouraging dietary-related information may facilitate patient participation in care and increase oral intakes among patients who have undergone colorectal surgery.

A systematic review of feeding practices among postoperative patients: is practice in-line with evidenced-based guidelines?

BACKGROUND: Early oral feeding after surgery is best practice among adult, noncritically ill patients. Evidenced-based guidelines (EBG) recommend commencing liquid and solid feeding within 24 h of surgery to improve patient (e.g. reduced morbidity) and hospital (e.g. reduced length of stay) outcomes. Whether these EBG are adhered to in usual clinical practice remains unknown. The present study aimed to identify the time to commencement of first oral feed (liquid or solid) and first solid feed among postoperative, noncritically ill, adult patients. METHODS: MEDLINE, CINAHL, SCOPUS and Web of Science databases were searched from inception to June 2016 for observational studies reporting liquid and/or solid feeding practices among postoperative patients. Studies reporting a mean/median time to first feed or first solid feed within 24 h of surgery or where ≥75% of patients were feeding by postoperative day one were considered in-line with EBG. RESULTS: Of 5826 articles retrieved, 29 studies were included. Only 40% and 22% of studies reported time to first feed and time to first solid feed in-line with EBG, respectively. Clear and free liquids were the first diet types commenced in 86% of studies. When solids were commenced, 44% of studies reported using various therapeutic diet types (e.g. light) prior to the commencement of a regular diet. Patients who underwent gastrointestinal procedures appeared more likely to experience delayed postoperative feeding. CONCLUSIONS: Our findings demonstrate a gap between postoperative feeding evidence and its practical application. This information provides a strong rationale for interventions targeting improved nutritional care following surgery.

Statistical mechanics of learning multiple orthogonal signals: asymptotic theory and fluctuation effects.

The learning of signal directions in high-dimensional data through orthogonal decomposition or principal component analysis (PCA) has many important applications in physics and engineering disciplines, e.g., wireless communication, information theory, and econophysics. The accuracy of the orthogonal decomposition can be studied using mean-field theory. Previous analysis of data produced from a model with a single signal direction has predicted a retarded learning phase transition below which learning is not possible, i.e., if the signal is too weak or the data set is too small then it is impossible to learn anything about the signal direction or magnitude. In this contribution we show that the result can be generalized to the case where there are multiple signal directions. Each nondegenerate signal is associated with a retarded learning transition. However, fluctuations around the mean-field solution lead to large finite size effects unless the signal strengths are very well separated. We evaluate the one-loop contribution to the mean-field theory, which shows that signal directions are indistinguishable from one another if their corresponding population eigenvalues are separated by O(N(-tau)) with exponent tau>1/3, where N is the data dimension. Numerical simulations are consistent with the analysis and show that finite size effects can persist even for very large data sets.

Rat brain serotonin neurones that express neuronal nitric oxide synthase have increased sensitivity to the substituted amphetamine serotonin toxins 3,4-methylenedioxymethamphetamine and p-chloroamphetamine.

Substituted amphetamines such as p-chloroamphetamine and the abused drug methylenedioxymethamphetamine cause selective destruction of serotonin axons in rats, by unknown mechanisms. Since some serotonin neurones also express neuronal nitric oxide synthase, which has been implicated in neurotoxicity, the present study was undertaken to determine whether nitric oxide synthase expressing serotonin neurones are selectively vulnerable to methylenedioxymethamphetamine or p-chloroamphetamine. Using double-labeling immunocytochemistry and double in situ hybridization for nitric oxide synthase and the serotonin transporter, it was confirmed that about two thirds of serotonergic cell bodies in the dorsal raphé nucleus expressed nitric oxide synthase, however few if any serotonin transporter immunoreactive axons in striatum expressed nitric oxide synthase at detectable levels. Methylenedioxymethamphetamine (30 mg/kg) or p-chloroamphetamine (2 x 10 mg/kg) was administered to Sprague-Dawley rats, and 7 days after drug administration there were modest decreases in the levels of serotonin transporter protein in frontal cortex, and striatum using Western blotting, even though axonal loss could be clearly seen by immunostaining. p-Chloroamphetamine or methylenedioxymethamphetamine administration did not alter the level of nitric oxide synthase in striatum or frontal cortex, determined by Western blotting. Analysis of serotonin neuronal cell bodies 7 days after p-chloroamphetamine treatment, revealed a net down-regulation of serotonin transporter mRNA levels, and a profound change in expression of nitric oxide synthase, with 33% of serotonin transporter mRNA positive cells containing nitric oxide synthase mRNA, compared with 65% in control animals. Altogether these results support the hypothesis that serotonin neurones which express nitric oxide synthase are most vulnerable to substituted amphetamine toxicity, supporting the concept that the selective vulnerability of serotonin neurones has a molecular basis.

Is there nicotinic modulation of nerve growth factor? Implications for cholinergic therapies in Alzheimer's disease.

Studies on the neurobiology of nerve growth factor (NGF) reveal a diverse range of actions. Through alterations in gene expression, NGF is important in maintaining and regulating the phenotype of neurons that express the high-affinity receptor, trkA. Nerve growth factor also has a rapid action, revealed by its role in pain signaling in bladder and in skin. In the central nervous system (CNS), NGF has an intimate relationship with the cholinergic system. It promotes cholinergic neuron survival after experimental injury but also maintains and regulates the phenotype of uninjured cholinergic neurons. In addition to these effects mediated by gene expression, NGF has a rapid neurotransmitter-like action to regulate cholinergic neurotransmission and neuronal excitability. Consistent with its actions on the cholinergic system, NGF can enhance function in animals with cholinergic lesions and has been proposed to be useful in humans with Alzheimer's disease (AD); however, the problems of CNS delivery and of side effects (particularly pain) limit the clinical efficacy of NGF. Drug treatment strategies to enhance production of NGF in the CNS may be useful in the treatment of AD. Nicotine is one such agent, which, when administered directly to the hippocampus in rats, produces long-lasting elevation of NGF production.

TrkA immunoreactive neurones in the rat spinal cord.

We report the presence in rat spinal cord of a novel neuronal system expressing tyrosine kinase receptor (trkA), the high affinity receptor for nerve growth factor (NGF). TrkA immunoreactive cell bodies were observed in the intermediate grey matter of the spinal cord and were classified into three main groups: central canal cells located dorsolateral to the aqueduct, partition cells located between lamina X, and the lateral border of the intermediate grey, and a morphologically heterogeneous group which included large cells located near the lateral border. In situ hybridization confirmed that cells in all these areas express trkA mRNA. Combined immunofluorescence and retrograde Fluoro-Gold labelling was used to further characterise the projections and neurotransmitter profile of the trkA cells. Although often located in the vicinity of preganglionic cell groups, trkA immunoreactive cells are not themselves preganglionic. Rather, the central canal and partition cells belong to a neurochemically complex cholinergic propriospinal system. Many partition cells coexpress trkA, choline acetyltransferase (ChAT), the low affinity neurotrophin receptor, p75, and nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d). In contrast, trkA immunoreactive central canal cells express ChAT, but do not express p75 and only a subpopulation express NADPH-d. The large trkA immunoreactive cells located on the lateral border do not express ChAT. TrkA immunoreactive fibres were also present and were located in the dorsal horn, in the dorsal columns, and in a bundle ventral to the aqueduct. However, double labelling revealed that the trkA immunoreactive fibres are not intrinsic but are primary afferent in origin and coexpress p75. The location of this novel trkA neuronal system is consistent with it having a role in the segmental integration of autonomic outflow. NGF could affect this system by modulating neuronal phenotype and/or synaptic efficacy.

Distribution of messenger RNAs encoding enkephalin, substance P, somatostatin, galanin, vasoactive intestinal polypeptide, neuropeptide Y, and calcitonin gene-related peptide in the midbrain periaqueductal grey in the rat.

The midbrain periaqueductal grey matter (PAG) has numerous functional roles that include mediating nociceptive inhibition and integrating behavioural and physiological responses to potentially threatening or stressful stimuli. Underlying these behaviours is the diverse interconnectivity of this region, and it is possible that neurochemical subdivisions within the PAG reflect the functional properties of the different PAG regions. In this study, using in situ hybridization, we have investigated the distribution in the rat PAG of the messenger ribonucleic acids (mRNAs) encoding seven neuropeptides: enkephalin (ENK), substance P (SP), somatostatin (SST), galanin (GAL), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), and calcitonin gene-related peptide (CGRP). Each peptide mRNA had a distinct topographical distribution in the PAG. Preproenkephalin A (ENK) mRNA-expressing cells were found at all levels of the PAG in three distinct longitudinal columns. Preprotachykinin A (SP)-expressing cells were found at all levels of the PAG, principally in the Edinger-Westphal nucleus and the lateral and dorsal PAG. There was a column of neurons producing mRNA-encoding somatostatin that extended along the rostrocaudal extent of the ventrolateral PAG; there were also labelled cells in the dorsal and dorsolateral subdivisions at some levels of the PAG. Galanin mRNA-producing neurones were limited to the dorsal raphe nucleus and to a second population in the ventral border of the aqueduct. VIP mRNA-producing neurones were found in very localized regions of the PAG, including the cell-sparse region immediately ventral to the aqueduct and the ventral part of the dorsal raphe nucleus. NPY mRNA-producing neurones were localized mainly in some cells of the Edinger-Westphal nucleus and dorsal raphe nucleus. CGRP mRNA-expressing neurons were limited to the oculomotor and trochlear nucleus. The results showed a topographical distribution of neuropeptides over the rostrocaudal extent of the PAG that is compatible with the emerging theory that the anatomical and functional specificity of the PAG is expressed in the form of longitudinally arranged neuronal columns that extend for varying distances along the rostrocaudal axis of the midbrain PAG.

Intraperitoneal insulin is more potent than subcutaneous insulin at restoring hepatic insulin-like growth factor-I mRNA levels in the diabetic rat: a functional role for the portal vascular link.

There is evidence that the hormonal control of hepatic IGF-I production is mediated by GH and insulin. To elucidate the role of these hormones further we administered s.c. or i.p. insulin (at 2.5 and 5.0 IU/day) and/or GH (0.8 IU/day) to rats made diabetic with streptozotocin 16 days previously. Hepatic IGF-I production was then assessed by quantifying hepatic IGF-I mRNA levels by autoradiography of Northern blots. Diabetes resulted in a fivefold reduction in hepatic IGF-I mRNA levels (optical density (OD) of the 0.7-1.1 kb band: controls, 1.3 +/- 0.09; diabetics, 0.28 +/- 0.08; P < 0.01), which was not significantly changed by treatment with s.c. insulin (OD: low dose, 0.55 +/- 0.05; high dose, 0.58 +/- 0.05) or low dose i.p. insulin (OD: 0.40 +/- 0.03). High dose i.p. insulin enhanced hepatic IGF-I mRNA levels (OD: 0.93 +/- 0.23) compared with diabetic rats (P < 0.01) and those given high dose s.c. insulin (P < 0.04), despite the blood glucose values being similar in the treated groups (i.p., 4.72 +/- 0.29 mmol/l; s.c., 3.32 +/- 0.03 mmol/l). Administration of GH alone partially restored the hepatic IGF-I mRNA level (OD: GH-treated, 1.00 +/- 0.05; diabetic, 0.28 +/- 0.08; P < 0.01), whilst having no effect on blood glucose values (diabetic, 36.35 +/- 0.45 mmol/l; GH-treated, 38.65 +/- 2.39 mmol/l). Additional administration of s.c. insulin completely restored IGF-I mRNA levels to those of controls (OD: low dose, 1.35 +/- 0.14; high dose, 1.27 +/- 0.18).(ABSTRACT TRUNCATED AT 250 WORDS)

Intraregional variation in expression of serotonin transporter messenger RNA by 5-hydroxytryptamine neurons.

The distribution of the messenger RNA encoding the 5-hydroxytryptamine transporter was investigated in rat brain. 5-Hydroxytryptamine transporter messenger RNA was found exclusively in the B1-B9 cell groups containing the cell bodies of 5-hydroxytryptamine neurons. Combined in situ hybridization and 5-hydroxytryptamine immunocytochemistry demonstrated 5-hydroxytryptamine transporter gene expression in the majority of and exclusively in 5-hydroxytryptamine neurons. Cells differed in their levels of expression of 5-hydroxytryptamine transporter messenger RNA and 5-hydroxytryptamine immunofluorescence, but with a tight correlation between the two parameters. Image analysis of cells from B7, the dorsal raphe nucleus, and B8, the median raphe nucleus, revealed significant differences between groups in the mean cellular level of 5-hydroxytryptamine transporter gene expression. Cells in the ventromedial subdivision of B7 displayed higher levels of expression than cells in B8 or cells in the lateral wings of B7. There was also heterogeneity in the distribution of the cellular levels of expression for two other genes expressed by 5-hydroxytryptamine neurons: l-aromatic amino acid decarboxylase messenger RNA and tryptophan hydroxylase messenger RNA. However, the relative levels of expression of these two genes within the four regions studied differed from that of 5-hydroxytryptamine transporter messenger RNA. These results indicate intraregional differences between 5-hydroxytryptamine neurons with respect to 5-hydroxytryptamine transporter messenger RNA levels. Such differences may account for the differential sensitivity of 5-hydroxytryptamine neurons to cytotoxins.

Reduction of GABA and glutamate transporter messenger RNAs in the severe-seizure genetically epilepsy-prone rat.

The genetically epilepsy-prone rat is an animal model of inherited generalised tonic-clonic epilepsy that shows abnormal susceptibility to audiogenic seizures and a lowered threshold to a variety of seizure-inducing stimuli. Recent studies suggest a crucial role for glutamate and GABA transporters in epileptogenesis and seizure propagation. The present study examines the levels of expression of the messenger RNAs encoding the glial and neuronal glutamate transporters, GLT-1 and EAAC-1, and the neuronal GABA transporter, GAT-1, in paired male genetically epileptic-prone rats and Sprague Dawley control rats using the technique of in situ hybridization. In a parallel study, semiquantitative immunoblotting was used to assess GLT-1 and EAAC-1 protein levels in similarly paired animals. Animals were assessed for susceptibility to audiogenic seizures on six occasions, and killed seven days following the last audiogenic stimulus exposure. Rat brains were processed for in situ hybridization with radioactive 35S-labelled oligonucleotide probes (EAAC-1 and GAT-1), 35S-labelled riboprobes (GLT-1), and Fluorescein-labelled riboprobes (GLT-1 and GAT-1) or processed for immunoblotting using subtype-specific antibodies for GLT-1 and EAAC-1. Semiquantitative analyses were carried out on X-ray film autoradiograms in several brain regions for both in situ hybridization and immunoblotting studies. Reductions in GAT-1 messenger RNA were found in genetically epileptic-prone rats in all brain regions examined (-8 to -24% compared to control). Similar reductions in GLT-1 messenger RNA expression levels were seen in cortex, striatum, and CA1 (-8 to -12%) of genetically epileptic-prone rats; the largest reduction observed was in the inferior colliculus (-20%). There was a tendency for a reduced expression of EAAC-1 messenger RNA in most regions of the genetically epileptic-prone rat brain although this reached statistical significance only in the striatum (-12%). In contrast, no significant differences in GLT-1 and EAAC-1 protein between genetically epileptic-prone rats and control animals were observed in any region examined, although there was a tendency to follow the changes seen with the corresponding messenger RNAs. These results show differences in the messenger RNA expression levels of three crucial amino acid transporters. For the two glutamate transporters, GLT-1 and EAAC-1, differences in messenger RNA levels are not reflected or are only partially reflected in the expression of the corresponding proteins.

Identification of 5-hydroxytryptamine receptors positively coupled to adenylyl cyclase in rat cultured astrocytes.

1. 5-Hydroxytryptamine (5-HT) elicited a dose-dependent stimulation of intracellular adenosine 3': 5'-cyclic monophosphate (cyclic AMP) accumulation in cultured astrocytes derived from neonatal rat (Sprague Dawley) thalamic/hypothalamic area with a potency (pEC50) of 6.68 +/- 0.08 (mean +/- s.e. mean). 2. In order to characterize the 5-HT receptor responsible for the cyclic AMP accumulation the effects of a variety of compounds were investigated on basal cyclic AMP levels (agonists) and 5-carboxamidotryptamine (5-CT) stimulated cyclic AMP levels (antagonists). The rank order of potency for the agonists investigated was 5-CT (pEC50 = 7.81 +/- 0.09) > 5-methoxytryptamine (5-MeOT) (pEC50 = 6.86 +/- 0.36) > 5-HT (pEC50 = 6.68 +/- 0.08). The following compounds, at concentrations up to 10 microM, did not affect basal cyclic AMP levels 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), cisapride, sumatriptan, DOI and RU 24969. The rank order of potency of antagonists was methiothepin (pKi = 7.98 +/- 0.25) > mesulergine (pKi = 7.58 +/- 0.18) > ritanserin (pKi = 7.20 +/- 0.24) > clozapine (pKi = 7.03 +/- 0.19) > mianserin (pKi = 6.41 +/- 0.19). The following compounds, at concentrations up to 10 microM, were inactive: ketanserin, WAY100635, GR127935. This pharmacological profile is consistent with that of 5-HT7 receptor subtype-mediated effects. 3. The cultured astrocytes exhibited regional heterogeneity in the magnitude of cyclic AMP accumulation (Emax). Cells cultured from the thalamic/hypothalamic area had significantly higher Emax values (588 +/- 75% and 572 +/- 63% of basal levels for 5-CT and 5-HT, respectively) compared to brainstem (274 +/- 51% and 318 +/- 46%, respectively) and colliculus astrocytes (244 +/- 15% and 301 +/- 24%, respectively). No significant differences in pEC50 (for either 5-HT or 5-CT) values were observed. 4. Reverse transcriptase-polymerase chain reaction (RT-PCR) with primers specific for the 5-HT7 receptor confirmed expression of messenger RNA for this receptor subtype by the cultured astrocytes derived from all regions investigated. Primers specific for the 5-HT6 receptor also amplified a cDNA fragment from the same samples. 5. From these findings, we conclude that astrocytes cultured from a number of brain regions express functional 5-HT receptors positively coupled to adenylyl cyclase and that the level of receptor expression or the efficiency of receptor coupling is regionally-dependent. The pharmacological profile of the receptor on thalamic/hypothalamic astrocytes suggests that the 5-HT7 receptor is the dominant receptor that is functionally expressed even though astrocyte cultures have the capacity to express both 5-HT6 and 5-HT7 receptor messenger RNA.

Growth hormone increases IGF-I, collagen I and collagen III gene expression in dwarf rat skeletal muscle.

The effect of short-term treatment with biosynthetic growth hormone (GH) of male dwarf rats was studied in EDL and soleus muscles. In situ hybridisation revealed that in the untreated dwarf rat collagen I, collagen III and insulin-like growth factor-I (IGF-I) mRNA is mainly expressed by fibroblasts between the muscle fibre areas. Quantitative image analysis showed that, 8 h after a single GH injection, the level of mRNA for all three genes increased compared to the untreated dwarf animal. IGF-I mRNA levels were similar in normals and untreated dwarf rats but significantly increased 8 h after a single GH injection in EDL (P < 0.01) and soleus (P < 0.001). In untreated dwarf rats, collagen I and III gene expression was significantly less than in normal animals (P < 0.001). Collagen III gene expression also increased significantly 8 h after a single GH injection, in both muscles (P < 0.01). Collagen I gene expression showed significant increases 8 and 24 h after GH treatment in EDL (P < 0.01), although the increases seen in soleus did not reach significance. The effects of multiple GH injections (one, two or four) did not appear to be additive. The results of the time course studies are consistent with an intermediary role for IGF-I in the production of collagen in muscle.

Acute nicotine decreases, and chronic nicotine increases the expression of brain-derived neurotrophic factor mRNA in rat hippocampus.

Acute nicotine administration (0.5 mg/kg i.p.) significantly decreased BDNF mRNA levels in dentate gyrus, CA3 and CA1 subfields of the rat hippocampus 2 h and 24 h after administration. However, with 7 days nicotine treatment, tolerance developed to the inhibitory effect of nicotine on BDNF mRNA expression and there was a significant increase in BDNF expression 2 h after the final injection in the CA1 region. These data suggests that changes in expression of hippocampal BDNF may be involved in the behavioural effects of nicotine observed after acute and chronic treatment.

Ligand autoradiographic receptor screening: receptor cDNA expression in replicas of transfected COS cells.

In this paper we validate a methodology, ligand autoradiographic receptor screening (LARS), for detecting expression of full length receptor cDNAs in COS cells. The method involves transfection of COS cells with receptor cDNAs by spheroplast fusion, production of filter replicas of the cell fragments, ligand binding to the receptors expressed in the membranes, and autoradiographic detection of bound ligand. A beta-adrenergic receptor cDNA cloned into a eukaryotic expression vector reliably induces high levels of beta-adrenergic receptor expression in 2-12% of COS cell colonies transfected with this plasmid after experimental conditions are optimized. Receptor expression is monitored by autoradiographic detection of 125iodocyanopindolol binding to COS cell fragments immobilized on polyester filter replicas. Binding displays appropriate pharmacological properties. The number of high-density binding spots per filter depends on the fraction of the spheroplasts in the fusion mixture that contain the beta-adrenergic receptor cDNA. A 100-plate LARS experiment can assess receptor expression in more than 10(4) transfected colonies. Thus detection of receptor-encoding sequences present in libraries in proportions as low as 1 in 10(4) should be possible. This technique may therefore be useful in detecting expression of other receptor cDNAs for which selective radioligands are available.

Repeated administration of MDMA down-regulates preprocholecystokinin mRNA expression but not tyrosine hydroxylase mRNA expression in neurones of the rat substantia nigra.

The effect of repeated administration of 3,4-methylenedioxymethamphetamine (MDMA) on the expression of tyrosine hydroxylase and preprocholecystokinin (CCK) messenger RNAs in substantia nigra was examined by in situ hybridisation histochemistry. Sections hybridised with 35S-labelled oligonucleotides were subjected to computerised image analysis to determine the density of silver grains above positively labelled cells as an index of steady state mRNA levels. In the substantia nigra pars compacta, CCK mRNA levels were significantly reduced in drug-treated animals 24 h and at 2 weeks after the last dose of MDMA (10 mg/kg i.p., twice daily for 4 days). In the same animals, MDMA caused no change in the level of tyrosine hydroxylase mRNA in this brain region. The results show that MDMA can produce changes in dopamine neurones. Furthermore, since tyrosine hydroxylase and cholecystokinin are co-expressed in substantia nigra pars compacta, these results suggest that the expression of the tyrosine hydroxylase and CCK genes are regulated independently.

Altered expression of group I metabotropic glutamate receptors in the hippocampus of amygdala-kindled rats.

Kindling is a well documented model of acquired focal epilepsy and synaptic plasticity in the nervous system. Previous biochemical studies have indicated an increase in mGluR-mediated phosphoinositide hydrolysis in the amygdala or hippocampus of fully kindled animals. In this study we have used in situ hybridisation techniques to examine the mRNA expression of group I metabotropic glutamate receptors (mGluR1 and mGluR5 both linked to phosphoinositide hydrolysis) in the hippocampus of amygdala-kindled animals sacrificed 24 h, 7 days or 28 days following the last electrically evoked stage 5 seizure, and in implanted non-stimulated control rats. Results indicate an initial up-regulation in mGluR1 mRNA (expressed as percentage of control) bilaterally in the DG (35-40%) and CA3 (16-48%), and unilaterally in CA4 (12%) in the 24 h post-kindled group. In kindled animals studied 7 days after the last seizure, these changes were either reduced or had returned to control levels. By 28 days mGluR1 mRNA levels had returned to control levels, with only a persistent increase in expression unilaterally in the DG (14%). In contrast, an initial down-regulation in mGluR5 mRNA was observed bilaterally in CA4 (-45 and -25%) and CA1 (-46 and -45%), and unilaterally in DG and CA3 (-27 and -42% respectively) 24 h after the last kindled seizure. In the 7 and 28 day kindled groups significant alterations in expression of mGluR5 mRNA were still apparent. These data show that the mRNAs for mGluR1 and mGluR5 are differentially regulated by kindling, indicating that the expression of each of these receptors is under independent regulatory control. These perturbations in mRNA expression may contribute to kindling epileptogenesis but are unlikely to account for the maintenance of the kindled state.

Hippocampal neurotrophin and trk receptor mRNA levels are altered by local administration of nicotine, carbachol and pilocarpine.

Cholinergic receptor agonists nicotine (nicotinic), carbachol (nicotinic/muscarinic) and pilocarpine (muscarinic) were administered into the hippocampus and mRNA levels of neurotrophins and their receptors determined using in situ hybridisation. Drug doses were carefully chosen to avoid the potentially confounding effects of seizure and cell death. Nicotine caused a long-lasting increase in nerve growth factor (NGF) mRNA in all subfields of the hippocampus. The increase was evident from 24 h up to 72 h after drug administration. This increase was dependent on excitatory amino acid neurotransmission as it was blocked by administration of an AMPA or NMDA receptor antagonist. In contrast, carbachol and pilocarpine produced a transient increase in NGF mRNA levels present 4-8 h after drug administration. Pilocarpine caused a transient increase in hippocampal brain-derived neurotrophic factor (BDNF) levels, with carbachol and nicotine showing the same trend. Nicotine and carbachol caused transient decreases in NT-3 mRNA levels in dentate gyrus and CA2 with pilocarpine showing a similar trend. Increases in mRNA encoding full-length trkB were seen 8 h after nicotine, with nicotine also causing elevations in a mRNA encoding a truncated isoform (trkB.T2). TrkC mRNA was not altered by any of the conditions used. The study suggests that muscarinic and nicotinic receptor activation in the hippocampus causes transient changes in all of the neurotrophins, but that NGF levels are selectively up-regulated by nicotinic receptor stimulation. The reciprocal interaction between NGF and ascending cholinergic systems may be a component of the cognitive enhancing effects of nicotine.

Variation in the expression of the mRNA for protein kinase C isoforms in the rat suprachiasmatic nuclei, caudate putamen and cerebral cortex.

Using in situ hybridization, we have examined mRNA expression for five isoforms of protein kinase C (PKC alpha, beta1, beta2, gamma and epsilon) in the rat suprachiasmatic nuclei (SCN) and other central site during the 24 h cycle. The signal for each of these isoforms shows a marked local density within the SCN. In the absence of photic cues, there are changes in the expression of the mRNAs for the four isoforms that are Ca2+-dependent (alpha, beta1, beta2 and gamma), but not for one of the Ca2+-independent PKCs (epsilon). PKC alpha mRNA exhibits a monophasic rhythm of expression in the SCN with a peak at early subjective night, circadian time (CT) 14. In contrast, the mRNAs for PKC beta1, beta2 and gamma show a biphasic rhythm in the SCN with peaks at early subjective day, CT 0, and early subjective night, CT 14. The four Ca2+-dependent isoforms may therefore subserve phase-related functions within the SCN at the onset of subjective night and, in the case of beta1, beta2 and gamma, also at the onset of subjective day. Variation in the mRNAs for PKC beta1 and gamma (but not for alpha, beta2 or epsilon) is also found in the caudate putamen and in the cingulate and parietal cortex; the biphasic pattern of expression for these mRNAs is precisely in phase with that observed in the SCN. The beta1 and gamma isoforms may therefore contribute to temporally regulated functions at sites outside the SCN. The present observations raise the possibility that receptor-mediated regulation of circadian functions is modulated or even gated by circadian changes in intracellular components that participate in distinct signal cascades.

Selective up-regulation of protein kinase C epsilon in granule cells after kainic acid-induced seizures in rat.

Kainate-induced seizure activity causes persistent changes in the hippocampus that include synaptic reorganization and functional changes in the mossy fibers. Using in situ hybridization histochemistry, the expression of PKC alpha, PKC beta, PKC gamma, PKC delta and PKC epsilon mRNAs was investigated in the hippocampus of adult rats following seizures induced by a s.c. injection of kainic acid. In CA1 and CA3, we found a significant decrease in PKC gamma mRNA 1 day after kainic acid which persisted for a 2nd day in CA1. None of the other PKC isoform mRNAs were altered in CA1 or CA3. In granule cells, a significant up-regulation specific to PKC epsilon mRNA was observed. One week after kainic acid administration, a marked increase in PKC epsilon immunoreactivity was found that persisted 2 months after kainic acid administration. PKC epsilon immunoreactivity was found associated with mossy fibers projecting to the hilus of the dentate gyrus and to the stratum lucidum of the CA3 field and presumably with the newly sprouted mossy fibers projecting to the supragranular layer. These data provide the first evidence for a long-lasting increase of the PKC epsilon in the axons of granule cells caused by kainate-induced seizures and suggest that PKC epsilon may be involved in the functional and/or structural modifications of granule cells that occur after limbic seizures.