%0 Journal Article
%~ PubMed
%A Bejjani, Charbel
%A Machaalani, Rita
%A Waters, Karen A
%T The dorsal motor nucleus of the vagus (DMNV) in sudden infant death syndrome (SIDS): Pathways leading to apoptosis.
%B Respiratory Physiology & Neurobiology
%D 2013
%C Netherlands
%I Elsevier BV
%V 185
%N 2
%P 203-210
%@ 1569-9048
%X 
%Z FOR Codes: 111403 110203


%0 Journal Article
%~ PubMed
%A Tang, Samantha
%A Machaalani, Rita
%A Waters, Karen A
%T Expression of brain-derived neurotrophic factor and TrkB receptor in the sudden infant death syndrome brainstem.
%B Respiratory Physiology & Neurobiology
%D 2012
%C Netherlands
%I Elsevier BV
%V 180
%N 1
%P 25-33
%@ 1569-9048
%X This study compared the expression of BDNF (proBDNF and rhBDNF forms) and its receptor TrkB, in the medulla of sudden infant death syndrome (SIDS) infants and infants who died from known causes (non-SIDS). This study also evaluated these markers in association with SIDS clinical risk factors including, sleep position, cigarette smoke exposure and gender. Brainstem tissue was immunohistochemically stained and quantitative analyses were made for eight nuclei of the caudal and rostral medulla. Compared to non-SIDS, SIDS infants had lower rhBDNF in the caudal nucleus of the solitary tract and higher TrkB in the caudal dorsal motor nucleus of the vagus. Within the SIDS cohort, prone sleep position was associated with lower rhBDNF in the caudal arcuate nucleus, and cigarette smoke exposure was associated with lower rhBDNF and TrkB in the inferior olivary nucleus. Abnormal expression of BDNF and TrkB suggests that neuroprotective functions of the BDNF/TrkB system may be reduced in respiratory-related nuclei of SIDS infants.
%Z FOR Codes: 110904


%0 Journal Article
%~ PubMed
%A Machaalani, Rita
%A Say, Meichien
%A Waters, Karen A
%T Effects of cigarette smoke exposure on nicotinic acetylcholine receptor subunits α7 and β2 in the sudden infant death syndrome (SIDS) brainstem.
%B Toxicology and applied pharmacology
%D 2011
%C United States
%I Academic Press
%V 257
%N 3
%P 396-404
%@ 1096-0333
%X It is postulated that nicotine, as the main neurotoxic constituent of cigarette smoke, influences SIDS risk through effects on nicotinic acetylcholine receptors (nAChRs) in brainstem nuclei that control respiration and arousal. This study compared ??7 and ??2 nAChR subunit expression in eight nuclei of the caudal and rostral medulla and seven nuclei of the pons between SIDS (n=46) and non-SIDS infants (n=14). Evaluation for associations with known SIDS risk factors included comparison according to whether infants had a history of exposure to cigarette smoke in the home, and stratification for sleep position and gender. Compared to non-SIDS infants, SIDS infants had significantly decreased ??7 in the caudal nucleus of the solitary tract (cNTS), gracile and cuneate nuclei, with decreased ??2 in the cNTS and increased ??2 in the facial. When considering only the SIDS cohort: 1-cigarette smoke exposure was associated with increased ??7 in the vestibular nucleus and increased ??2 in the rostral dorsal motor nucleus of the vagus, rNTS and Cuneate, 2-there was a gender interaction for ??7 in the gracile and cuneate, and ??2 in the cNTS and rostral arcuate nucleus, and 3-there was no effect of sleep position on ??7, but prone sleep was associated with decreased ??2 in three nuclei of the pons. In conclusion, SIDS infants demonstrate differences in expression of ??7 and ??2 nAChRs within brainstem nuclei that control respiration and arousal, which is independent on prior history of cigarette smoke exposure, especially for the NTS, with additional differences for smoke exposure (??2), gender (??7 and ??2) and sleep position (??2) evident.
%Z FOR Codes: 110903 111403


%0 Journal Article
%~ PubMed
%A Caldwell, Patrina
%A Hensley, Rebecca
%A Machaalani, Rita
%A Cheng, Alan
%A Waters, Karen
%T How effective is adenoidectomy alone for treatment of obstructive sleep apnoea in a child who presents with adenoid hypertrophy?
%B Journal of Paediatrics and Child Health
%D 2011
%C United Kingdom, Australia
%I Wiley-Blackwell Publishing Ltd.
%V 47
%N 8
%P 568-571
%@ 1034-4810
%X 
%Z FOR Codes: 111403


%0 Journal Article
%~ PubMed
%A Machaalani, Rita
%A Kashi, Petra K
%A Waters, Karen A
%T Distribution of nicotinic acetylcholine receptor subunits alpha7 and beta2 in the human brainstem and hippocampal formation.
%B Journal of chemical neuroanatomy
%D 2010
%C Netherlands
%I Elsevier BV
%V 40
%N 3
%P 223-31
%@ 1873-6300
%X We provide the first quantification and localization of the nicotinic acetylcholine receptors (nAChRs) alpha7 and beta2 in the human adult and infant brainstem and hippocampus. After applying immunohistochemistry on formalin fixed and paraffin embedded human brain tissue, we qualitatively analyzed the staining to provide a comparison in expression amongst the brainstem nuclei and hippocampal regions, and between the infants and adults. Amongst the brainstem regions studied, the greatest protein expression for both alpha7 and beta2 subunits was in the motor nuclei of the medulla and pons. Lowest expression for both subunits was in the midbrain nuclei, except for the oculomotor nucleus. Comparison between infants and adults showed greater expression in the infant brainstem nuclei of the dorsal motor nucleus of the vagus, hypoglossal, inferior olivary nucleus, nucleus of the solitary tract, abducens, and facial nuclei, for both alpha7 and beta2 nAChRs. In the hippocampus, only the alpha7 subunit showed greater expression in infants compared to adults. We conclude that both alpha7 and beta2 containing nAChRs play an important role in many nuclei of the human infant and adult brainstem, and that the alpha7 subunit may have a more prominent role in the infant than adult hippocampus.
%Z FOR Codes: 110903 111403 110399


%0 Journal Article
%~ PubMed
%A Machaalani, Rita
%A Gozal, Evelyne
%A Berger, François
%A Waters, Karen A
%A Dematteis, Maurice
%T Effects of Post-Mortem Intervals on Regional Brain Protein Profiles in Rats using SELDI-TOF MS Analysis.
%B Neurochemistry international
%D 2010
%C United Kingdom
%I Elsevier Ltd
%V 57
%N 6
%P 655-61
%@ 1872-9754
%X Identification of disease-associated proteins is critical for elucidating CNS disease mechanisms and elaborating novel treatment strategies. It requires post-mortem tissue analysis which can be significantly affected by the collection process, post-mortem intervals (PMIs), and storage conditions. To assess the effect of time and storage conditions on brain protein stability, SELDI-TOF-MS protein profiles were assessed in rat frontal cortex, caudate-putamen, hippocampus and medulla samples collected after various PMIs (0, 6, 12, 24, 48, and 72 h) at 4 ??C or at room temperature (RT) storage. Regions of interest were isolated from cryosections (tissue apposition, TA), or micropunched from cryosections apposed on filter paper (paper apposition, PA), and applied onto an NP20 ProteinChip array. Protein alterations, while greater at RT than at 4 ??C, were detected at 6h then differentially evolved in the various brain regions, with greater alterations in the caudate-putamen (60%) and the cortex (48%). Overall, our sensitive analytical method allowed unveiling of different patterns of protein susceptibility to PMI and to storage temperature in the various brain regions. Some protein peaks were altered in all brain regions and may potentially serve as markers of the PMI status of the brain, or for reference values when studying new proteins. Changes in disease-related proteins within post-mortem samples can be greatly affected by PMI and storage conditions, particularly when studying fragile and/or low abundant protein/peptides in tissues sampled from the caudate-putamen and neocortex.
%Z FOR Codes: 110999 601


%0 Journal Article
%~ PubMed
%A Tang, Samantha
%A Machaalani, Rita
%A Waters, Karen A
%T Immunolocalisation of pro- and mature- brain derived neurotrophic factor (BDNF) and receptor TrkB in the human brainstem and hippocampus.
%B Brain research
%D 2010
%C Netherlands
%I Elsevier BV
%V 1354
%N 
%P 1-14
%@ 0006-8993
%X Brain-derived neurotrophic factor (BDNF) and its receptor TrkB are essential in promoting normal development of the central nervous system. Specific functions that are affected in knockout models include respiratory control, coordination of movement and balance, and feeding activities. The expression of these markers has not yet been studied in the human infant brain. This study provides a detailed account of the distribution and localization of both pro- and mature-recombinant human (rh) forms of BDNF, and of TrkB in the human infant brainstem and hippocampus, and qualitatively compares this expression to that seen in the human adult. Using commercially available antibodies, we applied immunohistochemistry on formalin fixed and paraffin embedded human brain tissue [n=8 for infant, n=6 for adult], and qualitatively analyzed the expression of proBDNF, rhBDNF and TrkB. Amongst the brainstem regions studied, the greatest expression of the markers was in the mesencephalic trigeminal of the pons, and in the medulla, the inferior olive and arcuate nucleus. The lowest expression was in the substantia nigra of the midbrain and pontine locus coeruleus. Compared to adults, all the studied markers had a higher expression in the infant brainstem nuclei of the hypoglossal, vestibular, dorsal motor nucleus of the vagus, prepositus, cuneate, and dorsal raphe. In the hippocampus, only TrkB showed a higher expression in infants compared to adults. We conclude that BDNF and TrkB play important roles in controlling respiration, movement, balance and feeding in the brainstem and that the TrkB receptor is the most age-sensitive component of this system, especially in the hippocampus.
%Z FOR Codes: 110903


%0 Journal Article
%~ PubMed
%A Kashem, Mohammed A
%A Sarker, Ranjana
%A Des Etages, H
%A Machaalani, Rita
%A King, Nicholas
%A McGregor, Iain S
%A Matsumoto, Izuru
%T Comparative proteomics in the corpus callosal sub-regions of postmortem human brain.
%B Neurochemistry international
%D 2009
%C United Kingdom
%I Elsevier Ltd
%V 55
%N 7
%P 483-90
%@ 0197-0186
%X The corpus callosum (CC) is a single anatomical region with homologous cytoarchitecture and divided into four sub-regions such as the rostrum, the genu, the body and the splenium. Neuroimaging analysis revealed that susceptibility to clinical neurological diseases of these sub-regions is variable, indicating biochemical and physiological heterogenecity. To understand the biochemical make up of these regions, we compared the protein expression of these three sub-regional areas [the genu, the body and the splenium (n=9)] through 2D proteomics, which is a high-throughput global protein expression analysis technique. Normative proteomic comparison of gels, and analysis of spectra revealed that 17 (identified as 7 proteins), 35 (identified as 20 proteins) and 39 (identified as 21 proteins) protein spots were differentially expressed in the genu vs. the body, the genu vs. the splenium and the body vs. the splenium, respectively. These results suggest that the sub-regions of the CC differ at the level of protein expression. Identified proteins of the different groups belong to several functional classes such as cytoskeletal, metabolic, signaling, oxidative stress and calcium regulation. Interestingly, oxidative stress defense and glucose metabolic pathways of the splenium are quite different from the genu which might be correlated to region specific vulnerability of neuronal illness. Protein expression maps of these regions can be used as a reference source for future studies to investigate the molecular basis of functional differences and degree of pathogenesis of various neurodegenerative diseases of the CC.
%Z FOR Codes: 110903 30406 110106


%0 Journal Article
%~ PubMed
%A Tang, Samantha
%A Machaalani, Rita
%A Kashem, Mohammad
%A Matsumoto, Izuru
%A Waters, Karen
%T Intermittent Hypercapnic Hypoxia Induced Protein Changes in the Piglet Hippocampus Identified by MALDI-TOF-MS.
%B Neurochemical research
%D 2009
%C United States
%I Springer New York LLC
%V 34
%N 12
%P 2215-25
%@ 1573-6903
%X Intermittent hypercapnic hypoxia (IHH) induces protein changes in the brainstem, but its effects on the hippocampus have not yet been studied. Using a proteomics-based approach, we tested the hypothesis that IHH up-regulates apoptotic promoters and down-regulates apoptotic inhibitors in the developing hippocampus. Male piglets aged 13-14 days were assigned to control (n = 6) or IHH (n = 5) groups. Using two-dimensional polyacrylamide gel electrophoresis, matrix-assisted laser desorption/ionisation-time of flight-mass spectrometry (MALDI-TOF-MS), a total of 26 protein spots were differentially expressed in IHH compared to control group. Thirteen of these (6 up-regulated, 7 down-regulated) were identified including 14-3-3??/?? (increased), glial fibrillary acidic protein (increased) and a-internexin (decreased). Further analysis with western blot validated these proteins and immunohistochemistry showed specific regional changes in the subiculum, stratum radiatum and CA1 of the hippocampus. Most proteins identified were involved in promoting cell survival under apoptotic conditions. These findings improve our understanding of the cellular processes that occur in the hippocampus during IHH exposure, and have important implications in clinical settings where IHH is experienced, for example, during prone sleeping or with obstructive sleep apnea in an infant.
%Z FOR Codes: 110903


%0 Journal Article
%~ PubMed
%A Machaalani, Rita
%A Say, Meichien
%A Waters, Karen
%T Serotoninergic receptor 1A in the sudden infant death syndrome brainstem medulla and associations with clinical risk factors.
%B Acta neuropathologica
%D 2009
%C Germany
%I Springer
%V 117
%N 3
%P 257-65
%@ 1432-0533
%X The immunoreactivity of the serotoninergic receptor subtype 1A (5HT(1A)R) was quantitatively analyzed in the human infant brainstem medulla (caudal and rostral levels). We hypothesized that immunoreactivity of 5HT(1A)R would be reduced in infants diagnosed with sudden infant death syndrome (SIDS). In particular that those infants with known clinical risk factors (including cigarette smoke exposure, bed sharing and sleep position) would have greater changes than those without clinical risks. Comparing SIDS (n = 67) to infants who died suddenly with another diagnosis (non-SIDS, n = 25), we found decreased 5HT(1A)R immunoreactivity in the majority of the nuclei studied at the rostral medulla level including dorsal motor nucleus of the vagus (DMNV), nucleus of the solitary tract, vestibular, and inferior olivary nucleus (ION). There was a significant relationship with all risk factors for 5HT(1A)R, especially for DMNV, suggesting that 5HT(1A)Rs are highly vulnerable to various insults within the SIDS DMNV. This study not only provides further evidence of abnormalities within the brainstem serotoninergic system of SIDS infants, but also shows that these changes may be associated with exposure to clinical risk factors.
%Z FOR Codes: 110903


%0 Journal Article
%~ PubMed
%A Browne, Cherylea J
%A Sharma, Nidhi
%A Waters, Karen A
%A Machaalani, Rita
%T The effects of nicotine on the alpha-7 and beta-2 nicotinic acetycholine receptor subunits in the developing piglet brainstem.
%B International journal of developmental neuroscience : the official journal of the International Society for Developmental Neuroscience
%D 2009
%C United Kingdom, Cana
%I Pergamon
%V 28
%N 0
%P 1-7
%@ 1873-474X
%X Exposure to cigarette smoke is a major risk factor for sudden infant death syndrome (SIDS). We tested the hypothesis that nicotine increases expression of the nicotinic acetylcholine receptor (nAChR) subunits alpha7 and beta2 in a piglet model. Piglets exposed to 2mg/kg/day nicotine for 14 days postnatally (n=14) were compared to non-exposed controls (n=14), (equal gender proportions). Immunohistochemistry was performed to identify and quantify changes in, alpha7 and beta2 nAChR subunits in 8 nuclei of the medulla at both the rostral and caudal levels. Compared to controls, nicotine exposed piglets had decreased alpha7 in the rostral dorsal motor nucleus of the vagus (rDMNV) (p=0.01), and increased beta2 in the caudal DMNV (cDMNV) (p=0.05), caudal nucleus of the spinal trigeminal tract (cNSTT) (p=0.03) and caudal nucleus of the solitary tract (cNTS) (p=0.04). Analysis by gender showed that in the control group, compared to males, females had higher beta2 in the caudal hypoglossal (cXII) (p<0.01) and caudal inferior olivary (p=0.04) nuclei, while in the nicotine group females had higher beta2 in the cDMNV (p=0.02). Compared to control males, nicotine exposed males had lower beta2 in the cXII (p<0.01). Overall, changes in alpha7 were specific to nicotine exposure with no gender differentiation. Changes in beta2 were more widespread but showed gender-specific effects. These findings provide evidence that early postnatal exposure to nicotine significantly affects nAChR subunit expressions in the developing brainstem.
%Z FOR Codes: 110903 60603


%0 Journal Article
%~ PubMed
%A Tang, Samantha
%A Machaalani, Rita
%A Waters, Karen A
%T Brain-derived neurotrophic factor (BDNF) and TrkB in the piglet brainstem after post-natal nicotine and intermittent hypercapnic hypoxia.
%B Brain research
%D 2008
%C Netherlands
%I Elsevier BV
%V 1232
%N 1232
%P 195-205
%@ 0006-8993
%X Brain-derived neurotrophic factor (BDNF) and its receptor TrkB play a significant role in the regulation of cell growth, survival and death during central nervous system development. The expression of BDNF and TrkB is affected by noxious insults. Two insults during the early post-natal period that are of interest to our laboratory are exposure to nicotine and to intermittent hypercapnic hypoxia (IHH). Piglet models were used to mimic the conditions associated with the risk factors for the sudden infant death syndrome (SIDS) including post-natal cigarette smoke exposure (nicotine model) and prone sleeping where the infant is subjected to re-breathing of expired gases (IHH model). We aimed to determine the effects of nicotine and IHH, alone or in combination, on pro- and rhBDNF and TrkB expression in the developing piglet brainstem. Four piglet groups were studied, with equal gender ratios in each: control (n=14), nicotine (n=14), IHH (n=10) and nic+IHH (n=14). Applying immunohistochemistry, and studying six nuclei of the caudal medulla, we found that compared to controls, TrkB was the only protein significantly decreased after nicotine and nic+IHH exposure regardless of gender. For pro-BDNF and rhBDNF however, observed changes were more evident in males than females exposed to nicotine and nic+IHH. The implications of these findings are that a prior nicotine exposure makes the developing brainstem susceptible to greater changes in the neurotrophic effects of BDNF and its receptor TrkB in the face of a hypoxic insult, and that the effects are greater in males than females.
%Z FOR Codes: 110903


%0 Journal Article
%~ PubMed
%A Machaalani, Rita
%A Waters, Karen A
%T Neuronal cell death in the Sudden Infant Death Syndrome brainstem and associations with risk factors.
%B Brain
%D 2008
%C United Kingdom
%I Oxford University Press
%V 131
%N Pt 1
%P 218-228
%@ 1460-2156
%X Immunoreactive expression of three cell death markers was quantitatively analysed in the human infant brainstem medulla. We assessed active caspase-3, TUNEL and single-stranded DNA (ssDNA) in a cohort of 92 infants, and analysed for: (i) variations in the immunoreactive expression with development; (ii) comparison of infants diagnosed with the Sudden Infant Death Syndrome (SIDS, n = 67) to infants who died suddenly with another diagnosis (non-SIDS, n = 25); and (iii) correlations with known clinical risk factors for SIDS. Five nuclei from the brainstem medulla (caudal and rostral levels) were studied, including the hypoglossal (XII), dorsal motor nucleus of the vagus (DMNV), the dorsal column nuclei (gracile and cuneate) and the arcuate nucleus. Our main hypothesis was that neuronal cell death would be increased in SIDS compared to non-SIDS infants, and the increase would correlate with risk factors such as prone sleeping and cigarette smoke exposure. Comparing SIDS to non-SIDS, there was an increase in caspase-3 in the rostral DMNV (P = 0.01), and a trend to increased TUNEL in the arcuate nucleus (P = 0.1), which was statistically significant when comparing the male SIDS to male non-SIDS cohort (P = 0.04). No major changes for ssDNA immunoreactivity were found. Moreover, TUNEL expression was affected by post-conceptional age, by sleep-related risk factors (predominantly affecting the dorsal column nuclei), and by cigarette smoke exposure in the rostral DMNV and arcuate nucleus. Active caspase-3 was affected by post-conceptional age but only in the XII, while gender-related differences were seen in the arcuate nucleus. This study provides further evidence of increased apoptosis in the brainstem of SIDS infants, but shows for the first time that these changes are also affected by age and gender, and by clinical risk factors such as the sleep position and cigarette smoke exposure.
%Z FOR Codes: 110903 111403


%0 Journal Article
%~ PubMed
%A Machaalani, Rita
%A Makris, Angela
%A Thornton, Charlene
%A Hennessy, Annemarie
%T Vascular Endothelial Growth Factor Receptor 1 (Flt1) and Apoptosis in the Preeclamptic Placenta and Effects of in vivo Anti-hypertensive Exposure.
%B Hypertension in pregnancy : official journal of the International Society for the Study of Hypertension in Pregnancy
%D 2008
%C United Kingdom
%I Informa Healthcare
%V 27
%N 4
%P 361-73
%@ 1525-6065
%X Objective: To test the hypothesis that vascular endothelial growth factor receptor 1 (Flt1) is negatively correlated with apoptosis in preeclampsia placentae, and to examine the effects of antihypertensive medication on apoptosis. Methods: Flt1 and TUNEL immunoreactivity were quantitatively compared in the stromal decidual cells, villous trophoblasts, and endothelial cells of placentae from uncomplicated pregnancies (NP, n = 34) to those in patients with preeclampsia (PE, n = 30), and those in patients with preeclampsia with superimposed intrauterine growth restriction (PE + IUGR, n = 7). Further analyses determined any correlations with the antepartum use of the antihypertensives clonidine and hydralazine. Results: There was no difference in either Flt1 or TUNEL when comparing PE placentae (with or without IUGR) with NP. There were no correlations with the use of the antihypertensives. Conclusion. Apoptotic levels do not correlate with Flt1 in preeclampsia placentae and are not regulated by invivo exposure to the antihypertensives clonidine and hydralazine.
%Z FOR Codes: 111401


%0 Journal Article
%~ PubMed
%A Machaalani, Rita
%A Arlotto, Marie
%A Waters, Karen A
%A Gozal, Evelyne
%A Berger, François
%A Dematteis, Maurice
%T A Novel Method of Tissue Collection and Storage: Validation Using SELDI-TOF MS Analysis.
%B Clinical chemistry
%D 2007
%C United States
%I American Association of Clinical Chemistry
%V 53
%N 7
%P 1387-1389
%@ 0009-9147
%X 
%Z FOR Codes: 110106


%0 Journal Article
%~ PubMed
%A Machaalani, R
%A Rodriguez, M
%A Waters, K A
%T Active caspase-3 in the sudden infant death syndrome (SIDS) brainstem.
%B Acta neuropathologica
%D 2007
%C Germany
%I Springer
%V 113
%N 5
%P 577-584
%@ 0001-6322
%X In a retrospective postmortem study, we examined the neuronal expression of active caspase-3, a specific apoptotic marker, in the brainstem of 67 infants dying from sudden infant death syndrome (SIDS), and 25 age-matched control infants (non-SIDS). Neuronal immunostaining for active caspase-3 was semi-quantitatively scored in nuclei from five brainstem levels: rostral, mid and caudal pons, and rostral and caudal medulla. Regardless of the cause of death (SIDS vs. non-SIDS), age-related differences in active caspase-3 expression were identified, predominantly in the medulla. No gender-related differences were identified. Comparing SIDS to non-SIDS cases, increased active caspase-3 expression was restricted to four nuclei in the caudal pons (abducens, facial, superior olivary, and pontine nuclei) and two nuclei in the rostral medulla (hypoglossal and dorsal motor nucleus of the vagus). We conclude that neuronal apoptosis is increased in the brainstem of SIDS compared to non-SIDS infants.
%Z FOR Codes: 110903


%0 Journal Article
%~ PubMed
%A Say, Meichien
%A Machaalani, Rita
%A Waters, Karen A
%T Changes in serotoninergic receptors 1A and 2A in the piglet brainstem after intermittent hypercapnic hypoxia (IHH) and nicotine.
%B Brain research
%D 2007
%C Osney Mead, Oxford,
%I Elsevier Science
%V 1152
%N 
%P 17-26
%@ 0006-8993
%X We studied the effects of intermittent hypercapnic hypoxia (IHH) and/or nicotine on the immunoreactivity of serotoninergic (5-HT) receptors 1A and 2A in the piglet brainstem. These exposures were developed to mimic two common risk factors for Sudden Infant Death Syndrome (SIDS); prone sleeping (IHH) and cigarette smoke exposure (nicotine). Immunoreactivity for 5-HT(1A)R and 5-HT(2A)R were studied in four nuclei of the caudal medulla. Three exposure groups were compared to controls (n=14): IHH (n=10), nicotine (n=14), and nicotine+IHH (n=14). In control piglets, the immunoreactivity of 5-HT(1A)R was highest in the hypoglossal nucleus (XII), followed by inferior olivary nucleus (ION), nucleus of the solitary tract (NTS) and dorsal motor nucleus of the vagus (DMNV), whereas for 5-HT(2A)R, the immunoreactivity was highest in DMNV/NTS and then ION. Compared to controls, IHH reduced 5-HT(1A)R immunoreactivity in all studied nuclei (p<0.05) but had no effect on 5-HT(2A)R immunoreactivity. Nicotine reduced 5-HT(1A)R immunoreactivity in the DMNV, ION and NTS (p<0.001), and reduced 5-HT(2A)R immunoreactivity in DMNV/NTS (p<0.05). Nicotine+IHH reduced 5-HT(1A)R in DMNV, ION and NTS (p<0.001) but had no effect on 5-HT(2A)R immunoreactivity. Effects of nicotine on the DMNV were more significant in males compared to the females. These results show for the first time that IHH and/or nicotine can reduce 5-HT receptor immunoreactivity within functionally important nuclei of the piglet medulla. The findings support our hypothesis that 5-HT receptor abnormalities may be caused by postnatal exposures to clinically-relevant stimuli such as cigarette smoke exposure and/or prone sleeping.
%Z FOR Codes: 110903


%0 Journal Article
%~ PubMed
%A Machaalani, Rita
%A Radford, Jane L
%A Waters, Karen A
%T Tissue fixation effects on immunohistochemical staining of caspase-3 in brain tissue.
%B Applied immunohistochemistry & molecular morphology : AIMM / official publication of the Society for Applied Immunohistochemistry
%D 2007
%C United States
%I Lippincott Williams & Wilkins, Inc.
%V 15
%N 4
%P 463-470
%@ 1541-2016
%X Fixation methods for tissue often vary amongst clinical and research laboratories. To evaluate the effects of fixation method on studies of brain tissue, we examined immunohistochemical outcomes amongst 2 fixatives, 4 caspase-3 antibodies, and 2 species (human infants and piglets). Fixatives were 10% neutral buffered formalin (NBF) or 10% NBF and glacial acetic acid (FAA). Antibodies for caspase-3 were commercially obtained and included 2 for active caspase-3, and 2 for procaspase-3 (CASP3 and CPP32). Immunohistochemical staining of caspase-3 varied with fixation method, with the greatest effect of fixation method observed for the active caspase-3 antibodies and this effect was present in both species. In NBF-fixed tissue, active caspase-3 immunoreactivity was only visible microscopically, and was specific to neuronal cell bodies. In FAA-fixed tissue, active caspase-3 immunoreactivity was visible macroscopically, and predominantly present in fiber tracts and fasciculi compared with neuronal bodies. Fixation and species differences were also identified for the procaspase-3 antibodies, CASP3 and CPP32, where FAA-fixed pig tissue showed abundant staining of blood vessels that were not observed in the NBF-fixed pig tissue or in the human tissue. This study characterizes differences in immunohistochemical outcomes using commercially available antibodies for caspase-3, according to tissue fixation method and species.
%Z FOR Codes: 110999


%0 Journal Article
%~ PubMed
%A Fanous, A M
%A Machaalani, R
%A Waters, K A
%T N-methyl-d-aspartate receptor 1 changes in the piglet brainstem after nicotine and/or intermittent hypercapnic-hypoxia.
%B Neuroscience
%D 2006
%C 9650 Rockville Pike,
%I Pergamon-Elsevier Science Ltd
%V 142
%N 2
%P 401-9
%@ 0306-4522
%X Prone sleeping and cigarette smoke exposure are two major risk factors for the sudden infant death syndrome (SIDS). Utilizing piglet models of early postnatal nicotine and/or intermittent hypercapnic-hypoxia (IHH) exposure, we tested the hypothesis that these exposures, separately or combined, increase N-methyl-D-aspartate (NMDA) receptor 1 (NR1) expression in the brainstem medulla. We also tested for gender-specific effects. Three piglet exposure groups were compared against 14 controls; 1, nicotine [n = 14], 2, IHH [n = 10], and 3, nicotine+IHH [n = 14], with equal gender proportions in each group. Non-radioactive in situ hybridization and immunohistochemistry were performed for NR1 mRNA and protein expression, respectively, and were quantified in seven nuclei of the brainstem medulla. NR1 mRNA was significantly increased in the gracile and inferior olivary nucleus (ION) after nicotine exposure, in five of seven nuclei after IHH exposure, and in three of seven nuclei after nicotine+IHH. The increased mRNA changes were accompanied by increased protein only in the ION after IHH and nicotine+IHH (P = 0.019, and P = 0.008 respectively). By gender, control females had greater NR1 mRNA than males in the dorsal motor nucleus of vagus (P = 0.05) and for protein in the ION (P = 0.02). This gender difference was maintained after nicotine exposure in the ION with additional gender differences observed including greater mRNA in the cuneate nucleus (P = 0.04) and nucleus of the spinal trigeminal tract (P = 0.03) of males compared with females. Overall, more changes occurred at the mRNA level than protein, and IHH exposure induced more changes than nicotine or nicotine+IHH exposures. Together, these findings suggest that hypercapnic-hypoxic exposures (modeling prone sleeping or sleep apnea) are more likely to induce NMDA receptor changes in the developing brainstem than nicotine exposure alone.
%Z FOR Codes: 111799 111799


%0 Journal Article
%~ PubMed
%A Machaalani, R
%A Waters, K A
%T Postnatal nicotine and/or intermittent hypercapnic hypoxia effects on apoptotic markers in the developing piglet brainstem medulla.
%B Neuroscience
%D 2006
%C Netherlands
%I Elsevier BV
%V 142
%N 1
%P 107-17
%@ 0306-4522
%X The most important risk factors currently identified for the sudden infant death syndrome (SIDS) are prone sleeping and cigarette smoke exposure. In this study, we investigated the neuropathological sequelae of these risk factors by exposing piglets to intermittent hypercapnic-hypoxia (IHH) and/or nicotine (nic) in the early postnatal period. Our hypothesis was that either nic or IHH exposure could increase neuronal cell death, and that combined exposure (nic+IHH) would be additive. Four exposure patterns were studied: controls (n=14), IHH (n=10), nic (n=14), and nic+IHH (n=14). All groups had equal gender ratios. Nic exposure via an implanted osmotic minipump commenced within 48 h of birth and continued until age 13-14 days when animals were killed and brains collected. A total of 48 min of hypercapnic-hypoxia was delivered on the day immediately prior to killing in a pattern comprising 6 min of HH (8% O(2), 7% CO(2), balance N(2)) alternating with 6 min of air. Immunohistochemistry was performed to identify neurons positive for active caspase-3 and DNA fragmentation (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling, TUNEL) in seven nuclei of the caudal medulla. Staining quantification showed that: 1. IHH induced neuronal death (increased both TUNEL and casapse-3) in more brainstem nuclei than nicotine. 2. Females were more severely affected by IHH than males. 3. Where IHH and nicotine were combined, TUNEL expression was approximately 5% less than IHH alone, but changes in caspase-3 were variable. We conclude that acute exposure to IHH in the postnatal period is more neurotoxic than exposure to nicotine alone. Combined exposure to IHH and nicotine produced variable responses with some results suggesting that nicotine can be neuroprotective. These results indicate that environmental insults attributable to prone sleeping can produce neurotoxic sequelae in SIDS, with some regional specificity in the response. However, no consistent relationship is evident when combining the two insults.
%Z FOR Codes: 111403