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.

A randomised controlled trial of fluid restriction compared to oesophageal Doppler-guided goal-directed fluid therapy in elective major colorectal surgery within an Enhanced Recovery After Surgery program.

There is continued controversy regarding the benefits of goal-directed fluid therapy, with earlier studies showing marked improvement in morbidity and length-of-stay that have not been replicated more recently. The aim of this study was to compare patient outcomes in elective colorectal surgery patients having goal-directed versus restrictive fluid therapy. Inclusion criteria included suitability for an Enhanced Recovery After Surgery care pathway and patients with an American Society of Anesthesiologists Physical Status score of 1 to 3. Patients were intraoperatively randomised to either restrictive or Doppler-guided goal-directed fluid therapy. The primary outcome was length-of-stay; secondary outcomes included complication rate, change in haemodynamic variables and fluid volumes. Compared to restrictive therapy, goal-directed therapy resulted in a greater volume of intraoperative fluid, 2115 (interquartile range 1350 to 2560) ml versus 1500 (1200 to 2000) ml, P=0.008, and was associated with an increase in Doppler-derived stroke volume index from beginning to end of surgery, 43.7 (16.3) to 54.2 (21.1) ml/m(2), P <0.001, in the latter group. Length-of-stay was similar, 6.5 (5 to 9) versus 6 (4 to 9) days, P=0.421. The number of patients with any complication (minor or major) was similar; 0% (30) versus 52% (26), P=0.42, or major complications, 1 (2%) versus 4 (8%), P=0.36, respectively. The increased perioperative fluid volumes and increased stroke volumes at the end of surgery in patients receiving goal-directed therapy did not translate to a significant difference in length-of-stay and we did not observe a difference in the number of patients experiencing minor or major complications.

A paediatric palliative care programme in development: trends in referral and location of death.

PURPOSE: To describe the formation of a paediatric palliative care programme providing care in hospital, at home or in hospice, ensuring continuity of care where the child and family desire. STUDY DESIGN: Descriptive analysis was performed on referral source, diagnosis and reason for discharge for patients referred to the Palliative Care Team at the Children's Hospital of Eastern Ontario in Ottawa, Ontario, Canada from 1999 to 2007. RESULTS: 341 children were referred. 24% had a neurological condition, 21% had genetic disorders or congenital anomalies, 20% had cancer, 18% had metabolic or neurodegenerative conditions and 17% had another diagnosis. The major sources of referral included paediatricians, neonatologists, oncologists and intensivists. 55% of the children have died. 58% of these died in hospital, 27% at home and 15% in hospice. Of the remaining 152 children, 7% were discharged from the programme after clinical improvement, 4% were moved to another geographic location or an adult centre, 2% were not eligible, 1% declined services and 4% were lost to follow-up. The remaining 90 children continue to be followed-up. In the hospitalised patients who died, the annual referral rate increased from 20% to >50%. IMPLICATIONS: Referral to the palliative care team has increased over time in all diagnostic categories and from all sources. Most children died in hospital; however, a significant number of families chose end-of-life care at home or in a hospice.

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.

Principal-component-analysis eigenvalue spectra from data with symmetry-breaking structure.

Principal component analysis (PCA) is a ubiquitous method of multivariate statistics that focuses on the eigenvalues lambda and eigenvectors of the sample covariance matrix of a data set. We consider p, N-dimensional data vectors xi drawn from a distribution with covariance matrix C. We use the replica method to evaluate the expected eigenvalue distribution rho(lambda) as N--> infinity with p=alphaN for some fixed alpha. In contrast to existing studies we consider the case where C contains a number of symmetry-breaking directions, so that the sample data set contains some definite structure. Explicitly we set C=sigma2I+sigma(2)Sigma(S)(m=1)A(m)B(m)B(T)(m), with A(m)>0 for all m. We find that the bulk of the eigenvalues are distributed as for the case when the elements of xi are independent and identically distributed. With increasing alpha a series of phase transitions are observed, at alpha=A(-2)(m), m=1,2,..., S, each time a single delta function, delta(lambda-lambda(u)(A(m))), separates from the upper edge of the bulk distribution, where lambda(u)(A)=sigma(2)[1+A][1+(alphaA)(-1)]. We confirm the results of the replica analysis by studying the Stieltjes transform of rho(lambda). This suggests that the results obtained from the replica analysis are universal, irrespective of the distribution from which xi is drawn, provided the fourth moment of each element of xi exists.

Bayesian phylogenetics using an RNA substitution model applied to early mammalian evolution.

We study the phylogeny of the placental mammals using molecular data from all mitochondrial tRNAs and rRNAs of 54 species. We use probabilistic substitution models specific to evolution in base paired regions of RNA. A number of these models have been implemented in a new phylogenetic inference software package for carrying out maximum likelihood and Bayesian phylogenetic inferences. We describe our Bayesian phylogenetic method which uses a Markov chain Monte Carlo algorithm to provide samples from the posterior distribution of tree topologies. Our results show support for four primary mammalian clades, in agreement with recent studies of much larger data sets mainly comprising nuclear DNA. We discuss some issues arising when using Bayesian techniques on RNA sequence data.

Transgenic SOD1 G93A mice develop reduced GLT-1 in spinal cord without alterations in cerebrospinal fluid glutamate levels.

Glutamate-induced excitotoxicity is suggested to play a central role in the development of amyotrophic lateral sclerosis (ALS), although it is still unclear whether it represents a primary cause in the cascade leading to motor neurone death. We used western blotting, immunocytochemistry and in situ hybridization to examine the expression of GLT-1 in transgenic mice carrying a mutated (G93A) human copper-zinc superoxide dismutase (TgSOD1 G93A), which closely mimic the features of ALS. We observed a progressive decrease in the immunoreactivity of the glial glutamate transporter (GLT-1) in the ventral, but not in the dorsal, horn of lumbar spinal cord. This effect was specifically found in 14- and 18-week-old mice that had motor function impairment, motor neurone loss and reactive astrocytosis. No changes in GLT-1 were observed at 8 weeks of age, before the appearance of clinical symptoms. Decreases in GLT-1 were accompanied by increased glial fibrillary acidic protein (GFAP) levels and no change in the levels of GLAST, another glial glutamate transporter. The glutamate concentration in the cerebrospinal fluid (CSF) of TgSOD1 G93A mice was not modified at any of the time points examined, compared with age-matched controls. These findings indicate that the loss of GLT-1 protein in ALS mice selectively occurs in the areas affected by neurodegeneration and reactive astrocytosis and it is not associated with increases of glutamate levels in CSF. The lack of changes in GLT-1 at the presymptomatic stage suggests that glial glutamate transporter reduction is not a primary event leading to motor neurone loss.

Cumulant dynamics of a population under multiplicative selection, mutation, and drift.

We revisit the classical population genetics model of a population evolving under multiplicative selection, mutation, and drift. The number of beneficial alleles in a multilocus system can be considered a trait under exponential selection. Equations of motion are derived for the cumulants of the trait distribution in the diffusion limit and under the assumption of linkage equilibrium. Because of the additive nature of cumulants, this reduces to the problem of determining equations of motion for the expected allele distribution cumulants at each locus. The cumulant equations form an infinite dimensional linear system and in an authored appendix Adam Prügel-Bennett provides a closed form expression for these equations. We derive approximate solutions which are shown to describe the dynamics well for a broad range of parameters. In particular, we introduce two approximate analytical solutions: (1) Perturbation theory is used to solve the dynamics for weak selection and arbitrary mutation rate. The resulting expansion for the system's eigenvalues reduces to the known diffusion theory results for the limiting cases with either mutation or selection absent. (2) For low mutation rates we observe a separation of time-scales between the slowest mode and the rest which allows us to develop an approximate analytical solution for the dominant slow mode. The solution is consistent with the perturbation theory result and provides a good approximation for much stronger selection intensities.

Elevated levels of group-III metabotropic glutamate receptors in the inferior colliculus of genetically epilepsy-prone rats following intracollicular administration of L-serine-O-phosphate.

The selective group-III metabotropic glutamate receptor agonist, L-serine-O-phosphate (L-SOP), when injected bilaterally into the inferior colliculus of the sound sensitive genetically epilepsy-prone (GEP) rats produces a short proconvulsant excitation followed by a long phase of protection against sound-induced seizures lasting for 2-4 days. We have studied this prolonged suppression of audiogenic seizures using pharmacological and molecular biological approaches including semiquantitative RT-PCR and western blotting. The intracerebroventricular injection of the protein synthesis inhibitor cycloheximide (120 microg) 30 min beforehand significantly reduces the proconvulsant seizure activity and the prolonged anticonvulsant effect of intracollicular L-SOP (500 nmol/side). The sensitive semiquantitative RT-PCR revealed a significant up-regulation in mGlu(4) and mGlu(7) mRNA levels in the inferior colliculus at 2 days (maximum suppression of audiogenic seizures) after intracollicular L-SOP injection compared with the non-injected, 2-day post-vehicle treated and 7-day (return to expressing audiogenic seizures) post-drug or vehicle-treated groups. No significant changes were observed in mGlu(6) or mGlu(8) mRNA expression levels in drug-treated compared with control groups. Examination of mGlu(4a) and mGlu(7a) protein levels using western blotting showed a significant increase in mGlu(7a) but no significant change in mGlu(4a) protein levels 2 days after L-SOP treatment compared with the control groups (non-injected and 2-day vehicle-injected group). These results suggest that up-regulation of mGlu(7) receptors is involved in the prolonged anticonvulsant effect of L-SOP against sound-induced seizures in GEP rats. The potential use of mGlu(7) agonists as novel anti-epileptic agents merits investigation.

Expression of amyloid precursor protein, tau and presenilin RNAs in rat hippocampus following deafferentation lesions.

In this study, entorhinal cortex lesions and/or medial septal area cholinergic lesions were used in the rat to mimic some of the principal and earliest affects in Alzheimer's disease, namely hippocampal deafferentation. We wished to test the hypothesis that deafferentation lesions cause changes in the regulation of three proteins that are known to be important in Alzheimer's disease pathology, namely amyloid precursor protein, presenilin and tau. Expression of amyloid precursor protein mRNA was increased in several subfields of hippocampus when examined 1 week after entorhinal cortex lesion, but was reduced, compared to sham operated controls, after medial septal area cholinergic lesions. Cholinergic lesions were combined with entorhinal cortex lesions and produced no change in APP mRNA levels compared to controls. No significant changes were observed in the parietal cortex after entorhinal cortex or cholinergic lesions either alone or in combination. Tau mRNA level in hippocampus was unchanged after lesions. Presenilin-1 mRNA was expressed in the hippocampus at very low levels, and appeared to be increased following entorhinal cortex lesion. Our results support the hypothesis that amyloid precursor protein expression in hippocampal neurons is differentially affected by glutamatergic and cholinergic afferent input, and that presenilin-1, but not tau, may be subject to the same type of control in vivo.

Nicotine regulates 5-HT(1A) receptor gene expression in the cerebral cortex and dorsal hippocampus.

The 5-HT(1A) receptor has previously been shown to be important in mediating the behavioural effects of nicotine. It is possible that nicotine administration might regulate the levels of 5-HT receptors in limbic and cortical regions, and such regulations may underlie adaptive responses to nicotine in the central nervous system. The effects of acute and chronic systemic (--)-nicotine administration on 5-HT(1A) receptor gene expression were measured by in situ hybridization, in the rat cerebral cortex, dorsal hippocampus and lateral septum. In the cortex, acute nicotine (0.5 mg/kg i.p.) significantly increased the expression of 5-HT(1A) receptor mRNA 2 h and 24 h after injection. Similarly, acute nicotine significantly increased 5-HT(1A) receptor mRNA in the dentate gyrus (DG), CA3 and CA1 regions of the dorsal hippocampus 2 h and 24 h after injection. Acute nicotine was without effect in the lateral septum. Chronic nicotine (0.5 mg/kg i.p; twice daily for 7 days) significantly decreased 5-HT(1A) receptor mRNA in the cortex 2 h after the final injection, but was without effect at 24 h or 72 h. Chronic nicotine caused no changes in 5-HT(1A) mRNA in the lateral septum or dorsal hippocampus. These data demonstrate that nicotine regulates 5-HT(1A) receptor gene expression in the cortex and hippocampus. The rapid regulation of expression of 5-HT(1A) receptor mRNA leads to the hypothesis that nicotine-induced 5-HT release may alter the postsynaptic sensitivity to 5-HT.

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.

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.

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.

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.

Cultured astrocytes express messenger RNA for multiple serotonin receptor subtypes, without functional coupling of 5-HT1 receptor subtypes to adenylyl cyclase.

The literature describing the expression of 5-HT receptor subtypes by astrocytes is controversial and incomplete. It is clear that primary cultures of astrocytes express receptors of the 5-HT2 family coupled to phospholipase C and of the 5-HT7 receptor family positively coupled to adenylyl cyclase. Cultured astrocytes have also been reported to express receptors of the 5-HT1 family, although the exact subtypes present are unknown. In the present study we have investigated which of the known rat G-protein coupled 5-HT receptor mRNAs are expressed by cultured astrocytes. Reverse transcriptase-polymerase chain reaction (RT-PCR) revealed expression of 5-HT1A, 5-HT1B, 5-HT1D, 5-HT1F, 5-HT2A, 5-HT2B, 5-HT2C, 5-HT5B, 5-HT6 and 5-HT7 receptor mRNAs in astrocytes derived from 2-day old rats and cultured for 10-12 days. Messenger RNAs for 5-HT4 and 5-HT5A receptors were not detected. The functional expression of 5-HT1 receptor subtypes was investigated by measuring the ability of 5-HT1 receptor agonists: 8-OH-DPAT (5-HT1A receptors), RU24969 (5-HT1A, 5-HT1B, 5-HT1D, and 5-HT1F receptors) or sumatriptan (5-HT1B, 5-HT1D, and 5-HT1F receptors) to modulate forskolin or isoproterenol stimulated cAMP production. These compounds, at concentrations up to 10 microM, did not significantly attenuate cAMP production. These results indicate that although astrocytes express mRNA for each of the five 5-HT1 receptor subtypes which have been isolated from the rat, these receptors are not coupled to the inhibition of adenylyl cyclase.

Serotonin transporters in adult rat brain astrocytes revealed by [3H]5-HT uptake into glial plasmalemmal vesicles.

Cultured astrocytes derived from neonatal rat brain exhibited high affinity, Na+-dependent, paroxetine and fluoxetine sensitive [3H]5-HT uptake. Reverse transcriptase-PCR demonstrated that astrocytes in culture expressed messenger RNA for the cloned serotonin transporter protein which has been characterised as the neuronal serotonin transporter. Although the serotonin transporter in cultured astrocytes displayed a Km value approximately 10 times greater than found in adult brain synaptosomes, these observations indicated that astrocytes in vitro may express the same serotonin transporter as neurons. Reverse transcriptase-PCR demonstrated the presence of serotonin transporter mRNA in the adult rat cerebral cortex, suggesting that astrocytes in vivo may express low levels of this mRNA. To investigate whether astrocytes in the adult CNS express functional serotonin transporters, glial plasmalemmal vesicles were prepared from cerebral cortex, representing a subcellular fraction composed primarily of vesicles derived from astrocytes. These vesicles were characterised by [3H]-glutamate and [3H]-dopamine uptake and by immunoblot analysis, using glial and synaptic markers: glutamate synthase, SNAP-25 and synaptobrevin. [3H]5-HT was taken up into glial plasmalemmal vesicles in a high affinity (Km approximately 40 nM), Na+ dependent, paroxetine-sensitive manner. The [3H]5-HT uptake capacity (Vmax) in these vesicles was approximately one quarter of that observed in synaptosomes. These data indicate that astrocytes in culture and in vivo are capable of 5-HT uptake via the previously characterised 'neuronal' serotonin transporter.