Innervation of the vasa nervorum: changes in human diabetics.

Transperineurial and epineurial vessels are innervated by plexuses of unmyelinated axons. Human sural nerve biopsies were examined ultrastructurally and immunocytochemically with an antibody which recognizes a neuronal and neuroendocrine protein, PGP 9.5, to characterize perivascular axons of these plexuses. Diabetics exhibited a greater degree of abnormal innervation of the vasa nervorum than nondiabetics with and without neuropathy. Abnormal innervation included: a reduction in the percentage of vessels exhibiting perivascular axons and a concomitant increase in the percentage of vessels having denervated Schwann cell units, particularly around vessels confined to perineurial compartments, and remaining axons in nerves from diabetics exhibited fewer varicosities. Denervated arterioles of diabetics also displayed structural changes indicating injury. The arteriolar structural defects and loss of neurogenic control of neural blood flow may lead to or aggravate endoneurial ischemia or hypoxia. The patchy, focal endoneurial fiber loss that is prominent in proximal nerves and associated with the distal myelinated fiber loss of some diabetic patients may be due in part to perivascular denervation of the vasa nervorum.

Transperineurial arterioles in human sural nerve.

The perineurial sheath of nerve fascicles is a protective cellular layer that separates the endoneurium from the epineurium. Transperineurial arterioles (TPA) connect the endoneurial capillary plexuses to the epineurial arterial nutrient supply. Transperineurial arterioles are defined as any arteriole that is confined to a perineurial cell compartment, which would include all arterioles within the perineurium proper or within perineurial sleeves in the epi- or endoneurium. In this study of biopsied human sural nerves, three-dimensional reconstruction of one micron sections and ultrastructural analysis of step serials, we find that TPA are confined in open-ended perineurial sleeves by which they pass from the epineurium through the perineurial sheath proper into the endoneurium. Most TPA are terminal arterioles as evidenced by size (10-25 microns), morphological characteristics, and the fact that they connect with capillaries. Transperineurial arterioles gradually lose their continuous muscle coat and become post-arteriolar capillaries (PAC). Vascular segments that emerge into the endoneurium from the perineurial sleeves are generally of the PAC variety. Transperineurial arterioles and post-arteriolar capillaries are often associated with a plexus of unmyelinated nerve fibers. Axon varicosities exhibit a variety of morphologically distinct vesicles including dense-cored and a diversity of agranular vesicles. These findings suggest that TPA play a role in the neurogenic control of endoneurial blood flow.

Unmyelinated nerve fiber estimation by immunocytochemistry. Correlation with electron microscopy.

Electron microscopy (EM) is currently required for quantitation of unmyelinated nerve fiber (UMNF) densities. Electron microscopy is time-consuming, costly, and generally only considers a fraction of an entire nerve. Anti-PGP 9.5, which recognizes a neuron-associated antigen, may be used in glutaraldehyde-fixed, paraffin-embedded human sural nerve biopsies to identify unmyelinated axons. In nerves counterstained with Luxol fast blue, the correlation between EM-obtained UMNF densities and paraffin-obtained UMNF densities was excellent (p < 0.0001). In addition, myelinated nerve fiber (MNF) densities estimated by the same method from paraffin-embedded nerve gave excellent correlation with traditional morphometric estimates (p = 0.0026). PGP 9.5 immunocytochemistry enables the detection of minute axons (< 0.5 microns) and multiple axons per Schwann cell subunit, but does not allow for the preparation of accurate fiber size histograms or the analysis of UMNF pathology. Whether used for UMNF quantitation or as a qualitative review of the UMNF population of a nerve, this method is quicker and less expensive than traditional EM methodologies.

BRL 46470 potently antagonizes neural responses activated by 5-HT3 receptors.

The effect of a novel 5-HT3 receptor antagonist, BRL 46470, has been studied on two electrophysiological models for 5-HT3 receptors: grease-gap recordings from rat isolated vagus nerve and whole-cell patch-clamp recordings from mouse neuroblastoma-rat glioma NG108-15 cells. Its action on the rat vagus nerve was compared to that of four other 5-HT3 receptor antagonists. On the rat vagus, BRL 46470 reduced the maximum depolarizing response to 5-HT in a concentration-dependent manner with an IC50 of 0.3-1.0 nM, but the EC50 for 5-HT was not appreciably affected. This action was similar to that of granisetron and ICS 205-930, but differed from that of GR38032F and (+)-tubocurarine which produced clear rightward shifts of the concentration-response curve to 5-HT. The 5-HT-induced fast inward current of voltage-clamped NG108-15 cells was also antagonized by 1 nM BRL 46470 in an insurmountable manner. In contrast to (+)-tubocurarine, the action of BRL 46470 on the rat vagus nerve and NG108-15 cells did not readily reverse on washing with antagonist-free medium. It is concluded that BRL 46470 is a potent, insurmountable 5-HT3 receptor antagonist on the rat vagus and NG108-15 cells.

Multiple 5-HT receptors in the guinea-pig superior cervical ganglion.

1. We have studied the pharmacology of the depolarization by 5-hydroxytryptamine (5-HT) of the guinea-pig isolated superior cervical ganglion (SCG) using the grease-gap technique. We studied the effects of selective and non-selective antagonists on the responses to 5-HT and other 5-HT receptor agonists. 2. We have extended the pharmacology of the 5-HT3 receptor in this preparation by studying the effects of granisetron, BRL 46470 and mianserin on the concentration-response curve (CRC) to 2-methyl-5-HT. As with other 5-HT3 receptor antagonists, these compounds exhibited a lower affinity for guinea-pig 5-HT3 receptors than for rat 5-HT3 receptors. 3. We have confirmed that low concentrations of 5-HT (< or = 1 microM) mediate ketanserin-sensitive responses and higher concentrations of 5-HT also recruit 5-HT3 receptors. The responses to low concentrations of 5-HT were antagonized by low concentrations of ketanserin, spiperone, mianserin, DOI and LSD indicating probably mediation by 5-HT2A receptors. At high concentrations, the hallucinogen, DOI, but not LSD, evoked a ketanserin-sensitive depolarization. 4. Although mianserin could bind to the 5-HT2A receptors in this preparation, we could not demonstrate a down-regulation of depolarizations evoked by these receptors after a 10 day oral treatment with mianserin (10 mg kg-1, daily). 5. 5-Carboxamidotryptamine (5-CT) evoked a prolonged depolarization. Although high concentrations of 5-CT (> or = microM) appeared to activate 5-HT2A receptors, lower concentrations of 5-CT evoked a response with a distinct pharmacology. After studying the action of 20 selective and non-selective 5-HT receptor ligands we believe that this response may be mediated by a novel receptor; but its pharmacology is closest to that of receptors in the 5-HT2 receptor family. Like 5-CT, 5-HT (3-300 microM) could evoke an LSD-sensitive response in the presence of the 5-HT2 receptor antagonist, ketanserin and the 5-HT3 receptor antagonist, tropisetron (all 1 microM). 6. We conclude that 5-HT activates three pharmacologically distinct receptors to depolarize the guinea-pig SCG. Low concentrations of 5-HT appear to activate 5-HT2A receptors. Higher concentrations of 5-HT also activate 5-HT3 receptors and a possible novel 5-HT receptor. The novel receptor could be a species homologue of a 5-HT2 receptor or an, as yet, unclassified 5-HT receptor.

Differential effects of electroconvulsive shock on the glutamate receptor mRNAs for NR2A, NR2B and mGluR5b.

We have studied the effects of single and repeated electroconvulsive shock (ECS) treatment on the mRNA levels of several glutamate receptors in the dentate gyrus and CA1 regions of the rat brain. In the dentate gyrus, such treatment elevated the mRNAs for the NMDA subunits NR2A and NR2B, but it reduced the mRNA for the metabotropic glutamate receptor mGlu5b. With the exception of NR2A, this effect was specific to the dentate gyrus. The changes in NR2B mRNA lasted the longest, but all changes had returned to control values after 48 h. The possible significance of such changes to the antidepressant effect of ECT is discussed.

Factors influencing demand for primary medical care in women aged 20-44 years: a preliminary report.

This paper reports on a study designed to relate certain social and psychological variables to demand for medical care in a random sample of female patients aged between 20 and 44 years, in a National Health Service General Practice in the United Kingdom. Women selected for study completed questionnaires on anxiety and on their social characteristics. They also completed a health diary for four weeks, and over 12 months their demand for general practitioner care was recorded. This paper summarizes some of the literature on the subject of utilization behaviour, describes the objectives and methodology, and gives some preliminary results suggesting associations between anxiety, perception of symptoms and demand for primary care.

Doctor diagnosis and maternal recall of lower respiratory illness.

Epidemiological studies of respiratory illness in childhood have used the mother's recall of her child's respiratory illness experience as a measure of the frequency with which these illnesses occur. This paper explores the relationship between the diagnosis by the doctor of lower respiratory illnesses and the recall by the mother of the occurrence of these illnesses in a cohort of children in the first year of life. While there is a poor relationship between doctor diagnosis and maternal recall of these illnesses, the group of children reported by their mothers to have had bronchitis or pneumonia in the first year of life appear to suffer more respiratory illness than those children whose mothers recall no such illnesses.

5-HT2B receptor mRNA in guinea pig superior cervical ganglion.

Primers for 5-HT2A, 5-HT2B and 5-HT2C receptor mRNAs were used in reverse transcriptase-linked polymerase chain reactions (RT-PCR) to determine the presence of these transcripts in the guinea pig superior cervical ganglion. This was done to help identify an as yet unknown 5-HT2-like receptor which, in addition to 5-HT2A receptors, mediates a slow depolarization of this preparation. PCR products corresponding to 5-HT2A and 5-HT2B, but not 5-HT2C, receptor mRNA could readily be detected. Subsequent sequence analysis of these products confirmed that the 5-HT2A band corresponded to part of the guinea pig 5-HT2A receptor and the 5-HT2B band probably represents a portion of the guinea pig 5-HT2B receptor. The latter sequence shares greater homology with an equivalent region of the human than the rat 5-HT2B receptor.

Developmental changes in brain indoles, serum tryptophan and other serum neutral amino acids in the rat.

The rates at which brain neurons synthesize and release serotonin depend in part on brain tryptophan concentrations; these, in turn, vary directly with serum (or plasma) tryptophan, and inversely with the serum concentrations of other large neutral amino acids (LNAA). Concentrations of serum tryptophan, LNAA and brain indoles were examined in samples drawn at noontime from rats aged 0-59 days. Developmental changes in serum tryptophan largely paralleled those in the tryptophan/LNAA ratio, and brain tryptophan concentrations. Brain serotonin and 5-hydroxyindole acetic acid (5-HIAA) levels also increased postnatally; the changes in 5-HIAA tended to parallel those in brain tryptophan while those in serotonin did not.

Pharmacology of the 5-hydroxytryptamine-induced depolarization of the ferret vagus nerve in vitro.

Grease-gap recordings revealed that 5-hydroxytryptamine (5-HT) depolarized the ferret vagus nerve (pEC50 = 4.9). This response was mimicked by 2-methyl-5-HT and 1-phenylbiguanide, but not by 5-carboxamidotryptamine. Paroxetine (1 microM) or ketamine (10 microM) did not potentiate the response. Ketanserin (1 microM) did not reduce the depolarization, but four 5-HT3 receptor antagonists did. It is concluded that 5-HT depolarizes the ferret vagus nerve via 5-HT3 receptors, but these receptors may differ pharmacologically from those in other species.

AS-5370 potently antagonizes 5-HT3 receptor-mediated responses on NG108-15 cells and on the rat vagus.

The action of a novel 5-HT3 receptor antagonist, AS-5370, has been studied on two electrophysiological models for 5-HT3 receptors: whole-cell patch-clamp recordings from mouse neuroblastoma-rat glioma (NG108-15) cells and grease-gap recordings from rat isolated vagus nerve. The 5-hydroxytryptamine (5-HT)-induced fast inward current of voltage-clamped NG108-15 cells was antagonized by 1 nM AS-5370 in an insurmountable manner. On the rat vagus, AS-5370 reduced the maximum depolarizing response to 5-HT in a concentration-dependent manner. The IC50 for AS-5370 on the rat vagus was 0.3-1.0 nM. The EC50 for 5-HT on the rat vagus was not appreciably affected by AS-5370. On the rat vagus, the (R) enantiomer of AS-5370 was about 30 times more potent than the (S) enantiomer. The antagonist action of AS-5370 on these two cell types was compared with that of (+)-tubocurarine. Unlike tubocurarine, the effect of AS-5370 on NG108-15 cells was not readily reversed during wash. On the rat vagus, tubocurarine antagonized in a competitive manner with an IC50 of 0.3-1.0 microM (pKb = 7.2). It is concluded that AS-5370 is a potent 5-HT3 receptor antagonist on both NG108-15 cells and the rat vagus, but it does not act in a competitive manner.

The relationship between breast and bottle feeding and respiratory illness in the first year of life.

The relationship between breast or bottle feeding and the incidence of bronchitis and pneumonia in the first year of life was examined in a birth cohort of nearly 2000 children born in Harrow, England, in 1963--65. Fewer episodes of acute bronchitis and pneumonia were reported in children who were breast-fed than in children who were bottle-fed. Firstborn children were more likely to be breast-fed than subsequent children. Mothers who smoked were less likely to breast-feed their babies. Although birth order and parental smoking have been shown to be associated with bronchitis and pneumonia in the same cohort, the association between feeding pattern and respiratory illness history persisted when these and other important factors were taken into account.

Stereoselective interaction of mianserin with 5-HT3 receptors.

The interaction of the enantiomers of mianserin with the 5-HT3 receptor was determined. Using [3H]granisetron binding, (-)-mianserin was more potent than (+)-mianserin (pKi 8.46 and 6.95, respectively). The enantiomers competitively antagonized the depolarizing effect of 5-hydroxytryptamine in the rat vagus nerve preparation (pKapp: (-)-mianserin 8.13, (+)-mianserin 6.58). This stereoselectivity was maintained in-vivo as determined using ex-vivo inhibition of [3H]granisetron binding. Therefore, in contrast to its enantiomeric selectivity for the 5-HT1C and 5-HT2 receptors, where the (+)-isomer is more potent, the enantiomeric selectivity of mianserin for the 5-HT3 receptor was reversed. This differential selectivity of the enantiomers of mianserin may be useful in elucidating its utility in anxiety states.

Treatment of experimental spinal cord injury in ferrets.

A variety of treatment modalities upon a compressive model of spinal cord injury in ferrets was tested. Several of the treatments seemed to increase the degree of anatomical and/or functional sparing, but only dexamethasone did so in a statistically significant manner.

CL115,347, an analogue of prostaglandin E2. Peripheral circulatory effects of single ascending doses administered transdermally in normal subjects and in patients with Raynaud's phenomenon.

The beneficial effects of intra-arterial or intravenous infusion of the prostanoid products of anachridonic acid, PGE and prostacyclin (PGI2) are well documented. More recently an analogue of PGE2, (CL 115,347, American Cynamid Co.) has become available. This substance is absorbed transdermally from a patch placed on the skin. In a placebo-controlled trial the vasodilatory effect of single incremental dosage, 500 mcg, 1000 mcg and 1500 mcg, was measured in a temperature-/humidity controlled laboratory in normal subjects and in patients with primary and secondary Raynaud's phenomenon. The optimal dosage proved to be 1000 mcg; the effect may last for 84 hours; higher dosage may be associated with a "steal" phenomenon.

Do postal questionnaires change GPs' workload and referral patterns?

OBJECTIVE: We aimed to determine changes in workload in general practice associated with the postal administration of a health needs questionnaire. METHOD: We carried out controlled before-and-after intervention study of the effects of delivering a postal questionnaire to assess needs for care for patients with arthropathies of the hip and knee, groin hernia and varicose veins, and to assess health service utilization, general health status and risk factors for cardiovascular disease. The setting was a seven-partner, fundholding, group practice in Avon. The subjects were patients registered with an NHS group practice situated in Backwell and Nailsea, Avon. The outcome measures were the frequency of consultation, home visits and night visits, reasons for consultation, referral to specialist agencies and patterns of prescribing. RESULTS: There was no significant difference between the study and control group in the year before and the year after the postal administration of the questionnaire with respect to changes in overall frequency of consultation, frequency of referral (including type of referral) and frequency of prescribing of non-steroidal anti-inflammatory drugs. In the study group there was a significant (P<0.05) reduction in the number of daytime home visits and prescriptions written for analgesics. Analysis of the records of those who had received a medical examination, in addition to a postal questionnaire, showed that there was no significant difference between the study and control group with respect to frequency of consultation, referral to outside agencies or items prescribed. CONCLUSION: Administration of a health needs questionnaire to patients registered with this general practice was not associated with an increase in consultation frequency or referral, or a change in prescribing patterns. No plausible explanation could be identified for the significant reduction in the number of home visits and prescriptions written for analgesics. It was concluded that these results were a statistical artefact. On the basis of the evidence from this study, GPs can be reassured that the administration of health needs questionnaires of the type used in this study will not result in any increase in workload or costs of care incurred by increased referrals to outside agencies or increased prescribing.