Effectiveness of a sports-based HIV prevention intervention in the Dominican Republic: a quasi-experimental study.

Previous observational and quasi-experimental studies in sub-Saharan Africa have suggested the effectiveness of youth-targeted HIV prevention interventions using sport as an educational tool. No studies have yet assessed the effect of similar programs in the Caribbean. A quasi-experimental trial was conducted to assess the effectiveness of a sports-based intervention in six migrant settlements in the Puerto Plata Province of the Dominican Republic. A total of 397 structured interviews were conducted with 140 adolescents prior to, immediately following, and four months following 10-hour interventions using the Grassroot Soccer curriculum. Interview responses were coded, aggregated into composite scores, and analyzed using logistic regression, adjusting for baseline differences as well as age, sex, community, and descent. At post-intervention, significant differences were observed between groups in HIV-related knowledge (adjOR = 13.02, 95% CI = 8.26, 20.52), reported attitudes (adjOR = 12.01, 95% CI = 7.61, 18.94), and reported communication (adjOR = 3.13, 95% CI = 1.91, 5.12). These differences remained significant at four-month follow-up, though declines in post-intervention knowledge were observed in the Intervention group while gains in knowledge and reported attitudes were observed in the Control group. Results suggest that this sports-based intervention could play a valuable role in HIV prevention efforts in the Caribbean, particularly those targeting early adolescents. Further evaluation of sports-based interventions should include indicators assessing behavioral and biological outcomes, longer-term follow-up, a larger sample, randomization of study participants, and strenuous efforts to minimize loss-to-follow-up.

Molecular biology of the vesicular ACh transporter.

The cholinergic synapse has long been a model for biochemical studies of neurotransmission. The molecules that are responsible for synaptic transmission are being identified rapidly. The vesicular transporter for ACh, which is responsible for the concentration of ACh within synaptic vesicles, has been characterized recently, both at the molecular and functional level. Definitive identification of the cloned gene involved genetics of Caenorhabditis elegans, the specialized Torpedo electromotor system, and expression in mammalian tissue culture. Comparison of the vesicular transporter for ACh with the vesicular transporters for monoamines demonstrates a new gene family. Gene mapping has demonstrated a unique relationship between the genes for the vesicular ACh transporter and for choline acetyltransferase.

Localization of the Na(+)-coupled neutral amino acid transporter 2 in the cerebral cortex.

We studied the distribution and cellular localization of Na(+)-coupled neutral amino acid transporter 2, a member of the system A family of amino acid transporters, in the rat and human cerebral cortex using immunocytochemical methods. Na(+)-coupled neutral amino acid transporter 2-positive neurons were pyramidal and non-pyramidal, and Na(+)-coupled neutral amino acid transporter 2/GABA double-labeling studies revealed that Na(+)-coupled neutral amino acid transporter 2 was highly expressed by GABAergic neurons. Double-labeling studies with the synaptophysin indicated that rare axon terminals express Na(+)-coupled neutral amino acid transporter 2. Na(+)-coupled neutral amino acid transporter 2-immunoreactivity was also found in astrocytes, leptomeninges, ependymal cells and choroid plexus. Electron microscopy showed robust Na(+)-coupled neutral amino acid transporter 2-immunoreactivity in the somato-dendritic compartment of neurons and in glial processes, but, as in the case of double-labeling studies, failed to reveal Na(+)-coupled neutral amino acid transporter 2-immunoreactivity in terminals. To rule out the possibility that the absence of Na(+)-coupled neutral amino acid transporter 1- and Na(+)-coupled neutral amino acid transporter 2-positive terminals was due to insufficient antigen detection, we evaluated Na(+)-coupled neutral amino acid transporter 1/synaptophysin and Na(+)-coupled neutral amino acid transporter 2/synaptophysin coexpression using non-standard immunocytochemical procedures and found that Na(+)-coupled neutral amino acid transporter 1 and Na(+)-coupled neutral amino acid transporter 2+ terminals were rare in all conditions. These findings indicate that Na(+)-coupled neutral amino acid transporter 1 and Na(+)-coupled neutral amino acid transporter 2 are virtually absent in cortical terminals, and suggest that they do not contribute significantly to replenishing the Glu and GABA transmitter pools through the glutamate-glutamine cycle. The strong expression of Na(+)-coupled neutral amino acid transporter 2 in the somato-dendritic compartment and in non-neuronal elements that are integral parts of the blood-brain and brain-cerebrospinal fluid barrier suggests that Na(+)-coupled neutral amino acid transporter 2 plays a role in regulating the levels of Gln and other amino acids in the metabolic compartment of cortical neurons.

Mortality associated with congenital heart defects in the United States: trends and racial disparities, 1979-1997.

BACKGROUND: Surgical series and some population-based studies have documented a decrease in mortality from heart defects. Recent population-based data for the United States are lacking, however. We examined population-based data for patterns, time trends, and racial differences of mortality from heart defects for the United States from 1979 through 1997. METHODS AND RESULTS: We examined the multiple-cause mortality files compiled by the National Center for Health Statistics of the CDC from all death certificates filed in the United STATES: From these data, we derived death rates (deaths per 100 000 population) by the decedent's age, race, year of death, and heart defect type. We also analyzed age at death as an indirect indicator of survival. From 1979 through 1997, mortality from heart defects (all ages) declined 39%, from 2.5 to 1.5 per 100 000 population; among infants, the decline was 39%, or 2.7% per year. In 1995 to 1997, heart defects contributed to 5822 deaths per year. Of these deaths, 51% were among infants and 7% among children 1 to 4 years old. Mortality was on average 19% higher among blacks than among whites; this gap does not appear to be closing. Age at death increased for most heart defects, although less among blacks than among whites. CONCLUSIONS: Mortality from heart defects is declining in the United States, although it remains a major cause of death in infancy and childhood. Age at death is increasing, suggesting that more affected persons are living to adolescence and adulthood. The racial discrepancies should be investigated to identify opportunities for prevention.

Differential responsiveness of the pituitary-thyroid axis to thyrotropin-releasing hormone in mouse lines selected to differ in central nervous system sensitivity to ethanol.

Long-sleep (LS) and short-sleep (SS) mice are genetic lines that differ in central nervous system sensitivity to ethanol. The possible role of TRH in mediating the difference in the thyroid status between these two lines was investigated. An increase in TRH gene expression in the paraventricular nucleus and TRH peptide levels in the hypothalamus between postnatal days 8-14 in both SS and LS mice coincided with increased circulating levels of thyroxine during this critical period of central nervous system development. No significant differences in TRH biosynthesis were observed between LS and SS mice during this time. Exogenous administration of TRH to LS and SS mice on day 8, when endogenous serum thyroxine levels were equivalent, resulted in a greater increase in serum thyroxine in SS mice (150%) than LS mice (51%). The differential response to the TRH stimulation test was also present on day 14 (SS, 43%; LS, 18%). The differential responsiveness of the pituitary-thyroid axis to exogenous TRH paralleled the differential increase in endogenous serum thyroxine observed between day 8 and 14 in these mice. Administration of TRH to day 20 and adult (60 days) LS and SS mice resulted in nearly equivalent (approximately 75%) increases in free thyroxine serum levels, yet the magnitude of thyroxine release was 50% greater in SS mice, due perhaps to between-line differences within the thyroid glands. It is unlikely that dissimilar endogenous levels of TRH account for the intrinsic difference in the thyroid status in LS and SS mice. Instead, the increased pituitary-thyroid responsiveness to TRH in SS mice during the second postnatal week may translate into increased functional capacity of the thyroid gland in adult SS relative to LS mice.

Vesicular neurotransmitter transporters. Potential sites for the regulation of synaptic function.

Neurotransmission depends on the regulated release of chemical transmitter molecules. This requires the packaging of these substances into the specialized secretory vesicles of neurons and neuroendocrine cells, a process mediated by specific vesicular transporters. The family of genes encoding the vesicular transporters for biogenic amines and acetylcholine have recently been cloned. Direct comparison of their transport characteristics and pharmacology provides information about vesicular transport bioenergetics, substrate feature recognition by each transporter, and the role of vesicular amine storage in the mechanism of action of psychopharmacologic and neurotoxic agents. Regulation of vesicular transport activity may affect levels of neurotransmitter available for neurosecretion and be an important site for the regulation of synaptic function. Gene knockout studies have determined vesicular transport function is critical for survival and have enabled further evaluation of the role of vesicular neurotransmitter transporters in behavior and neurotoxicity. Molecular analysis is beginning to reveal the sites involved in vesicular transporter function and the sites that determine substrate specificity. In addition, the molecular basis for the selective targeting of these transporters to specific vesicle populations and the biogenesis of monoaminergic and cholinergic synaptic vesicles are areas of research that are currently being explored. This information provides new insights into the pharmacology and physiology of biogenic amine and acetylcholine vesicular storage in cardiovascular, endocrine, and central nervous system function and has important implications for neurodegenerative disease.

Cloning and expression of the vesamicol binding protein from the marine ray Torpedo. Homology with the putative vesicular acetylcholine transporter UNC-17 from Caenorhabditis elegans.

Complementary DNA clones corresponding to a messenger RNA encoding a 56 kDa polypeptide have been obtained from Torpedo marmorata and Torpedo ocellata electric lobe libraries, by homology screening with a probe obtained from the putative acetylcholine transporter from the nematode Caenorhabditis elegans. The Torpedo proteins display approximately 50% overall identity to the C. elegans unc-17 protein and 43% identity to the two vesicle monoamine transporters (VMAT1 and VMAT2). This family of proteins is highly conserved within 12 domains which potentially span the vesicle membrane, with little similarity within the putative intraluminal glycosylated loop and at the N- and C-termini. The approximately 3.0 kb mRNA species is specifically expressed in the brain and highly enriched in the electric lobe of Torpedo. The Torpedo protein, expressed in CV-1 fibroblast cells, possesses a high-affinity binding site for vesamicol (Kd = 6 nM), a drug which blocks in vitro and in vivo acetylcholine accumulation in cholinergic vesicles.

Preservation of nucleus basalis neurons containing choline acetyltransferase and the vesicular acetylcholine transporter in the elderly with mild cognitive impairment and early Alzheimer's disease.

Immunocytochemistry for choline acetyltransferase (ChAT) and the vesicular acetylcholine transporter (VAChT) was used to examine the expression of these linked cholinergic markers in human basal forebrain, including cases with early stages of Alzheimer's disease (AD). Previous neurochemical studies have measured decreased ChAT activity in terminal fields, but little change or even increased levels of VAChT. To determine total cholinergic neuron numbers in the nucleus basalis of Meynert (nbM), stereologic methods were applied to tissue derived from three groups of individuals with varying levels of cognition: no cognitive impairment (NCI), mild cognitive impairment (MCI), and early-stage Alzheimer's disease (AD). Both markers were expressed robustly in nucleus basalis neurons and across all three groups. On average, there was no significant difference between the number of ChAT- (210,000) and VAChT- (174, 000) immunopositive neurons in the nbM per hemisphere in NCI cases for which the biological variation was calculated to be 17%. There was approximately a 15% nonsignificant reduction in the number of cholinergic neurons in the nbM in the AD cases with no decline in MCI cases. The number of ChAT- and VAChT-immunopositive neurons was shown to correlate significantly with the severity of dementia determined by scores on the Mini-Mental State Examination, but showed no relationship to apolipoprotein E allele status, age, gender, education, or postmortem interval when all clinical groups were combined or evaluated separately. These data suggest that cholinergic neurons, and the coexpression of ChAT and VAChT, are relatively preserved in early stages of AD.

Differential co-localisation of the P2X7 receptor subunit with vesicular glutamate transporters VGLUT1 and VGLUT2 in rat CNS.

Presynaptic P2X(7) receptors are thought to play a role in the modulation of transmitter release and have been localised to terminals with the location and morphology typical of excitatory boutons. To test the hypothesis that this receptor is preferentially associated with excitatory terminals we combined immunohistochemistry for the P2X(7) receptor subunit (P2X(7)R) with that for two vesicular glutamate transporters (VGLUT1 and VGLUT2) in the rat CNS. This confirmed that P2X(7)R immunoreactivity (IR) is present in glutamatergic terminals; however, whether it was co-localised with VGLUT1-IR or VGLUT2-IR depended on the CNS region examined. In the spinal cord, P2X(7)R-IR co-localised with VGLUT2-IR. In the brainstem, co-localisation of P2X(7)R-IR with VGLUT2-IR was widespread, but co-localisation with VGLUT1-IR was seen only in the external cuneate nucleus and spinocerebellar tract region of the ventral medulla. In the cerebellum, P2X(7)R-IR co-localised with both VGLUT1 and VGLUT2-IR in the granular layer. In the hippocampus it was co-localised only with VGLUT1-IR, including in the polymorphic layer of the dentate gyrus and the substantia radiatum of the CA3 region. In other forebrain areas, P2X(7)R-IR co-localised with VGLUT1-IR throughout the amygdala, caudate putamen, striatum, reticular thalamic nucleus and cortex and with VGLUT2-IR in the dorsal lateral geniculate nucleus, amygdala and hypothalamus. Dual labelling studies performed using markers for cholinergic, monoaminergic, GABAergic and glycinergic terminals indicated that in certain brainstem and spinal cord nuclei the P2X(7)R is also expressed by subpopulations of cholinergic and GABAergic/glycinergic terminals. These data support our previous hypothesis that the P2X(7)R may play a role in modulating glutamate release in functionally different systems throughout the CNS but further suggest a role in modulating release of inhibitory transmitters in some regions.

Pregnancy experience after delivery of a child with a major birth defect: a population study.

OBJECTIVE: Data from a large population-based, case-control study were analyzed to determine whether women giving birth to children with major birth defects have different subsequent pregnancy patterns than those giving birth to live-born babies without defects. Other studies examining this phenomenon have been smaller, have not been population-based, or have not addressed the different effects that a wide range of major defects might have on mothers' subsequent pregnancy rates. METHODS: Mothers of 4918 infants with major birth defects born from 1968 through 1980 in metropolitan Atlanta were compared with mothers of 3029 control infants, frequency-matched on birth year, birth hospital, and race. RESULTS: The pregnancy rate in the first 3 years after the index birth was higher among case mothers (36%) than among control mothers (30%, P < .0001). This excess was seen for mothers of stillborn case infants (64%) and mothers of case infants who survived the first year of life (31%). Pregnancy rates varied by birth defect type. Maternal and infant factors varied among case and control subjects and influenced subsequent pregnancy rates. CONCLUSION: The reproductive behavior observed in this study supports the theory that mothers of nonsurviving children with birth defects compensate by acting to "replace" the lost child. Reproductive behavior was also strongly associated with having completed a previous pregnancy and by the type of birth defect.

Periconceptional multivitamin use and the occurrence of conotruncal heart defects: results from a population-based, case-control study.

OBJECTIVE: The preventive efficacy of the periconceptional use of multivitamins is well established for neural tube defects, much less so for other birth defects. We conducted a population-based, case-control study to assess the effects of multivitamin use on the risk for conotruncal defects, a group of severe heart defects that includes transposition of the great arteries, tetralogy of Fallot, and truncus arteriosus. METHODS: From the population-based Atlanta Birth Defects Case-Control Study, we identified 158 case infants with conotruncal defects and 3026 unaffected, randomly chosen control infants, born from 1968 through 1980 to mothers residing in metropolitan Atlanta. Periconceptional multivitamin use was defined as reported regular use from 3 months before conception through the third month of pregnancy. We present the results of the crude analysis, because the multivariate model yielded essentially identical results. RESULTS: Mothers who reported periconceptional multivitamin use had a 43% lower risk of having infants with conotruncal defects (odds ratio [OR], 0.57; 95% confidence interval [CI], 0.33 to 1.00) than did mothers who reported no use. The estimated relative risk was lowest for isolated conotruncal defects (OR, 0.41; 95% CI, 0.20 to 0.84) compared with those associated with noncardiac defects (OR, 0.91; 95% CI, 0.33 to 2.52) or a recognized syndrome (OR, 1.82; 95% CI, 0.31 to 10.67). Among anatomic subgroups of defects, transposition of the great arteries showed the greatest reduction in risk (OR, 0.36; 95% CI, 0.15 to 0.89). CONCLUSIONS: Periconceptional multivitamin use is associated with a reduced risk for conotruncal defects. These findings could have major implications for the prevention of these birth defects.

Congenital malformations and intrauterine growth retardation: a population study.

The relationship between congenital malformations and intrauterine growth retardation was investigated using data from the population-based Metropolitan Atlanta Congenital Defects Program. Between 1970 and 1984, the system ascertained 13,074 infants with major structural malformations diagnosed in the first year of life and born to metropolitan Atlanta residents. These infants were classified as having intrauterine growth retardation if their birth weight was below the race-, sex-, and gestational age-specific tenth percentile limits for all Atlanta births. Overall, the frequency of intrauterine growth retardation among malformed infants was 22.3% (relative risk 2.6). Of 48 defect categories evaluated, 46 were associated with excess intrauterine growth retardation, most notably chromosomal anomalies (eg, 83.7% for infants with trisomy 18, relative risk 46) and anencephaly (73.3%, relative risk 25). Only a few isolated defects (such as isolated polydactyly, pyloric stenosis, and congenital hip dislocation) were not associated with excess intrauterine growth retardation. Among infants with multiple malformations, the frequency of intrauterine growth retardation increased markedly with increasing number of defects--from 20% for infants with two defects to 60% for infants with nine or more defects. The relationship between malformations and intrauterine growth retardation can be explained by one or more of three mechanisms: (1) intrauterine growth retardation can be a secondary disturbance to the presence of malformations; (2) intrauterine growth retardation can predispose the fetus to malformations; and (3) intrauterine growth retardation can coexist with malformations because of common etiologic factors.(ABSTRACT TRUNCATED AT 250 WORDS)

Clinical-epidemiologic assessment of pattern of birth defects associated with human teratogens: application to diabetic embryopathy.

The concepts of sensitivity, specificity, and predictive value can be used to assess patterns of birth defects associated with human teratogens. Although sensitivity of any single defect is generally low for many known teratogens, the presence of specific defect combinations is usually predictive of the teratogen. To evaluate the patterns of birth defects associated with diabetic embryopathy, a sensitivity-specificity analysis was performed on 4929 infants with major defects ascertained by the population-based Metropolitan Atlanta Congenital Defects Program between 1968 and 1980. By reviewing hospital records, maternal insulin-dependent diabetes mellitus was confirmed in 26 infants. Patterns of defects were evaluated among infants born to mothers with insulin-dependent diabetes mellitus and compared with the rest of the Metropolitan Atlanta Congenital Defects Program case population. Multiple logistic regression analysis was used to assess defect combinations that predict for insulin-dependent diabetes mellitus. Of 26 infants, 8 had multiple defects. However, most defects and their combinations were poorly sensitive and predictive for insulin-dependent diabetes mellitus. The predictive value for insulin-dependent diabetes mellitus was greatest for the combination of vertebral and cardiovascular anomalies (6.5%). Also, several pathogenetic mechanisms were noted among patients with insulin-dependent diabetes mellitus, such as cell migration defects, cell death events, deformations, and cardiac flow lesions. The inability to find a clear-cut phenotype for diabetic embryopathy may be due to several etiologic factors and mechanisms associated with diabetic embryopathy.

A population study of the VACTERL association: evidence for its etiologic heterogeneity.

Using the population-based data from the Metropolitan Atlanta Congenital Defects Program, the interrelation of the six defects that are components of the VACTERL association were investigated. There were 400 cases with two or more of these defects, whereas only 29 cases would be expected if the defects had occurred together randomly. There were 76 cases with three or more defects, whereas less than one case was expected. Of these 76 cases, seven had recognized causes (five chromosomal anomalies, two single-gene disorders); another 19 had recognized clinical phenotypes or syndromes of unknown etiology. In the remaining 50 cases, ventricular septal defect was the most common cardiovascular defect (30.0%), and renal agenesis was the most common renal anomaly (30%). Their most common limb defects were reduction deformities (34%) and polydactyly (20%). This study confirms the clinically recognized nonrandom occurrence of the VACTERL association. It also shows that the association is a spectrum of various combinations of its components, which can be a manifestation of several recognized disorders, rather than a distinct anatomic or etiologic entity. A common denominator of the VACTERL association is suggested to be a defective mesodermal development during embryogenesis, due to a variety of causes and leading to overlapping manifestations.

Diabetes mellitus during pregnancy and the risks for specific birth defects: a population-based case-control study.

Although the excess risk for birth defects among children of mothers with diabetes mellitus is well documented, there are few data concerning the risk for specific malformations. In the Atlanta Birth Defects Case-Control Study, those risks for malformations were evaluated. The population-based study included 4929 live and stillborn babies with major malformations ascertained by the Metropolitan Atlanta Congenital Defects Program in the first year of life born to residents of Metropolitan Atlanta between 1968 and 1980. The study also included 3029 nonmalformed live babies who were frequency-matched to case babies by race, period of birth, and hospital of birth. The relative risk for major malformations among infants of mothers with insulin-dependent diabetes mellitus (n = 28) was 7.9 (95% confidence interval [CI]1.9, 33.5) compared with infants of nondiabetic mothers. The relative risks for major central nervous system and cardiovascular system defects were 15.5 (95% CI = 3.3, 73.8) and 18.0 (95% CI = 3.9, 82.5), respectively. The absolute risks for major, central nervous system, and cardiovascular system malformations among infants of diabetic mothers were 18.4, 5.3, and 8.5 per 100 live births, respectively. Infants of mothers with gestational diabetes mellitus who required insulin during the third trimester of pregnancy were 20.6 (95% CI = 2.5, 168.5) times more likely to have major cardiovascular system defects than infants of nondiabetic mothers. The absolute risk for infants of this group of diabetic mothers was 9.7%. No statistically significant differences were found among infants of mothers with gestational diabetes mellitus who did not require insulin during pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)

A case-control study of maternal alcohol consumption and intrauterine growth retardation.

PURPOSE: Heavy maternal drinking during pregnancy causes fetal alcohol syndrome, but whether more moderate alcohol consumption is associated with such adverse pregnancy outcomes as intrauterine growth retardation (IUGR) remains controversial. METHODS: Using data from a case-control study, we examined the association between maternal alcohol consumption and risk for IUGR among 701 case and 336 control infants born during 1993-1995 in Monroe County, New York. RESULTS: Our results provide no evidence of an independent association between moderate maternal alcohol consumption (<14 drinks per week) and risk for IUGR. The risk for IUGR among heavy drinkers (> or =14 drinks per week) around the time of conception was OR = 1.4 (95% CI 0.7-2.6) for IUGR < or = 5th percentile and OR = 1.4 (95% CI 0.7-2.8) for IUGR 5th-10th percentile. For heavy drinkers during the first trimester, the OR was 1.3 (95% CI 0.4-4.5) for IUGR < or = 5th percentile and OR = 1.3 (95% CI 0.4-4.8) for IUGR 5th-10th percentile. CONCLUSIONS: Since IUGR is a heterogeneous outcome with a possible multifactorial origin, further studies are needed to examine the combined effects of alcohol and other environmental and genetic factors on IUGR risk for subgroups of IUGR.

Localization of vesicular monoamine transporter isoforms (VMAT1 and VMAT2) to endocrine cells and neurons in rat.

Polyclonal antipeptide antibodies have been raised against each of the two isoforms of the rat vesicular monoamine transporter, VMAT1 and VMAT2. Antibody specificity was determined by isoform-specific staining of monkey fibroblasts programmed to express either VMAT1 or VMAT2. The expression of VMAT1 and VMAT2 in the diffuse neuroendocrine system of the rat has been examined using these polyclonal antibodies specific for either VMAT1 or VMAT2. VMAT1 is expressed exclusively in endocrine/paracrine cells associated with the intestine, stomach, and sympathetic nervous system. VMAT2 is expressed in neurons of the sympathetic nervous system, and aminergic neurons in the enteric and central nervous systems. VMAT2 is expressed in at least two endocrine cell populations in addition to its expression in neurons. A subpopulation of chromogranin A (CGA)-expressing chromaffin cells of the adrenal medulla also express VMAT2, and the oxyntic mucosa of the stomach contains a prominent population of CGA- and VMAT2-positive endocrine cells. The expression of VMAT2 in neurons, and the mutually exclusive expression of VMAT1 and VMAT2 in endocrine/paracrine cell populations of stomach, intestine, and sympathetic nervous system may provide a marker for, and insight into, the ontogeny and monoamine-secreting capabilities of multiple neuroendocrine sublineages in the diffuse neuroendocrine system.

Human and monkey cholinergic neurons visualized in paraffin-embedded tissues by immunoreactivity for VAChT, the vesicular acetylcholine transporter.

The predicted C-terminal dodecapeptide of the human vesicular acetylcholine transporter (VAChT), deduced from the unique open reading frame of the recently cloned human VAChT cDNA, was conjugated through an N-terminal cysteine to keyhole limpet hemocyanin and used as an immunogen to generate polyclonal antihuman VAChT antibodies in rabbits. The distribution of the VAChT antigen in representative regions of the cholinergic nervous system was examined and compared to that of the acetylcholine biosynthetic enzyme choline acetyltransferase (ChAT), a specific marker for cholinergic neurons. VAChT immunoreactivity was localized in cell bodies of neurons in the basal forebrain and ventral horn of the spinal cord, regions in which major cholinergic projection systems to the cerebral cortex and to skeletal muscle, respectively, originate. The primate caudate nucleus contained numerous VAChT-positive interneurons. VAChT immunoreactivity was visualized in both cell bodies and extensive terminals in striatal interneurons, in contrast to formalin-fixed, deparaffinized sections stained for ChAT, in which cell bodies and fibers were stained but nerve terminals were less well visualized than with the VAChT antiserum. VAChT-positive nerve fibers were visualized in routinely immersion-fixed, paraffin-embedded human cerebral cortex, comparable to the density of fibers observed in perfusion-fixed Bouin's-postfixed monkey cerebral cortex. Extensive investment of virtually all principal ganglion cells of thoracic sympathetic ganglia of monkey and human with VAChT-positive nerve terminals was observed. VAChT-positive cell bodies, presumably corresponding to cholinergic sympathetic sudomotor neurons, were a significant fraction of the total principal cell population in monkey and human thoracic sympathetic ganglia.

Reserpine- and tetrabenazine-sensitive transport of (3)H-histamine by the neuronal isoform of the vesicular monoamine transporter.

The transport of (3)H-histamine by the endocrine-specific (VMAT1) and neuronal (VMAT2) isoforms of the vesicular monoamine transporter has been evaluated in digitonin-permeabilized fibroblasts transfected with either VMAT1 or VMAT2. Transport of (3)H-histamine by both VMAT1 and VMAT2 was reserpine-sensitive but only transport by VMAT2 was inhibited by tetrabenazine. Maximal equilibrated levels of (3)H-histamine accumulation by VMAT2 (K(m) 300 mu M) were approximately three times greater than that mediated by VMAT1 when using a subsaturating concentration of exogenous (3)H-histamine (50 mu M). The expression of VMAT2 in histaminergic neurons in the rat brain was examined with polyclonal antipeptide antibodies specific for VMAT1 or VMAT2. VMAT2-positive and tyrosine hydroxylase-negative immunoreactive cell bodies were localized to the ventral part of the posterior hypothalamus in the region of the mamillary nuclei. The transport properties of VMAT2 and the distribution of VMAT2 in cell bodies in the tuberomammillary nucleus of the posterior hypothalamus reported here and the apparent absence of VMAT1 and VMAT2 in tissue mast cells support previous findings of reserpine-sensitive and reserpine-resistant pools of histamine in brain and peripheral tissues.

Distribution of the vesicular acetylcholine transporter (VAChT) in the central and peripheral nervous systems of the rat.

Expression of the acetylcholine biosynthetic enzyme choline acetyltransferase (ChAT), the vesicular acetylcholine transporter (VAChT), and the high-affinity plasma membrane choline transporter uniquely defines the cholinergic phenotype in the mammalian central (CNS) and peripheral (PNS) nervous systems. The distribution of cells expressing the messenger RNA encoding the recently cloned VAChT in the rat CNS and PNS is described here. The pattern of expression of VAChT mRNA is consistent with anatomical, pharmacological, and histochemical information on the distribution of functional cholinergic neurons in the brain and peripheral tissues of the rat. VAChT mRNA-containing cells are present in brain areas, including neocortex and hypothalamus, in which the existence of cholinergic neurons has been the subject of debate. The demonstration that VAChT is a completely adequate marker for cholinergic neurons should allow the systematic delineation of cholinergic synapses in the rat nervous system when antibodies directed to this protein are available.