Do maternal opioids reduce neonatal regional brain volumes? A pilot study.

OBJECTIVE: A substantial number of children exposed to gestational opioids have neurodevelopmental, behavioral and cognitive problems. Opioids are not neuroteratogens but whether they affect the developing brain in more subtle ways (for example, volume loss) is unclear. We aimed to determine the feasibility of using magnetic resonance imaging (MRI) to assess volumetric changes in healthy opioid-exposed infants. STUDY DESIGN: Observational pilot cohort study conducted in two maternity hospitals in New South Wales, Australia. Maternal history and neonatal urine and meconium screens were obtained to confirm drug exposure. Volumetric analysis of MRI scans was performed with the ITK-snap program. RESULT: Scans for 16 infants (mean (s.d.) gestational age: 40.9 (1.5) weeks, birth weight: 3022.5 (476.6) g, head circumference (HC): 33.7 (1.5 cm)) were analyzed. Six (37.5%) infants had HC <25th percentile. Fourteen mothers used methadone, four used buprenorphine and 11 used more than one opioid (including heroin, seven). All scans were structurally normal whole brain volumes (357.4 (63.8)) and basal ganglia (14.5 (3.5)) ml were significantly smaller than population means (425.4 (4.8), 17.1 (4.4) ml, respectively) but lateral ventricular volumes (3.5 (1.8) ml) were larger than population values (2.1(1.5)) ml. CONCLUSION: Our pilot study suggests that brain volumes of opioid-exposed babies may be smaller than population means and that specific regions, for example, basal ganglia, that are involved in neurotransmission, may be particularly affected. Larger studies including correlation with neurodevelopmental outcomes are warranted to substantiate this finding.

Cannabis, the pregnant woman and her child: weeding out the myths.

To review and summarise the literature reporting on cannabis use within western communities with specific reference to patterns of use, the pharmacology of its major psychoactive compounds, including placental and fetal transfer, and the impact of maternal cannabis use on pregnancy, the newborn infant and the developing child. Review of published articles, governmental guidelines and data and book chapters. Although cannabis is one of the most widely used illegal drugs, there is limited data about the prevalence of cannabis use in pregnant women, and it is likely that reported rates of exposure are significantly underestimated. With much of the available literature focusing on the impact of other illicit drugs such as opioids and stimulants, the effects of cannabis use in pregnancy on the developing fetus remain uncertain. Current evidence indicates that cannabis use both during pregnancy and lactation, may adversely affect neurodevelopment, especially during periods of critical brain growth both in the developing fetal brain and during adolescent maturation, with impacts on neuropsychiatric, behavioural and executive functioning. These reported effects may influence future adult productivity and lifetime outcomes. Despite the widespread use of cannabis by young women, there is limited information available about the impact perinatal cannabis use on the developing fetus and child, particularly the effects of cannabis use while breast feeding. Women who are using cannabis while pregnant and breast feeding should be advised of what is known about the potential adverse effects on fetal growth and development and encouraged to either stop using or decrease their use. Long-term follow-up of exposed children is crucial as neurocognitive and behavioural problems may benefit from early intervention aimed to reduce future problems such as delinquency, depression and substance use.

Amphetamines, the pregnant woman and her children: a review.

The objective of this study is to review and summarize available evidence regarding the impact of amphetamines on pregnancy, the newborn infant and the child. Amphetamines are neurostimulants and neurotoxins that are some of the most widely abused illicit drugs in the world. Users are at high risk of psychiatric co-morbidities, and evidence suggests that perinatal amphetamine exposure is associated with poor pregnancy outcomes, but data is confounded by other adverse factors associated with drug-dependency. Data sources are Government data, published articles, conference abstracts and book chapters. The global incidence of perinatal amphetamine exposure is most likely severely underestimated but acknowledged to be increasing rapidly, whereas exposure to other drugs, for example, heroin, is decreasing. Mothers known to be using amphetamines are at high risk of psychiatric co-morbidity and poorer obstetric outcomes, but their infants may escape detection, because the signs of withdrawal are usually less pronounced than opiate-exposed infants. There is little evidence of amphetamine-induced neurotoxicity and long-term neurodevelopmental impact, as data is scarce and difficult to extricate from the influence of other factors associated with children living in households where one or more parent uses drugs in terms of poverty and neglect. Perinatal amphetamine-exposure is an increasing worldwide concern, but robust research, especially for childhood outcomes, remains scarce. We suggest that exposed children may be at risk of ongoing developmental and behavioral impediment, and recommend that efforts be made to improve early detection of perinatal exposure and to increase provision of early-intervention services for affected children and their families.

Coordinated outpatient care of the narcotic-dependent infant.

OBJECTIVE: Concern regarding parental capacity to manage their infant's drug withdrawal traditionally leads to prolonged hospitalization for both mother and infant. This study examines the impact of a multidisciplinary follow-up clinic for infants discharged home on morphine. METHODOLOGY: Records of full-term infants born to mothers with narcotic dependency were reviewed retrospectively. Two periods were compared: 1995-1997 (period A) and 1998 to September 1999 with clinic established (period B). RESULTS: Twenty-five and 26 mothers were identified in periods A and B, respectively. Almost half had fewer than four antenatal clinic visits and most were on methadone with other substance usage. Despite higher maternal methadone doses (mean 48.5 vs 89.5 mg/day, P = 0.009) and withdrawal rates, the mean length of stay was significantly shorter for period B mothers (7.8 +/- 4.8 vs 5.4 +/- 3.3 days, P = 0.01) and babies (14.8 +/- 9.7 vs 8.7 +/- 7.2, P = 0.01). Median duration of morphine treatment was significantly shorter in period A (17 vs 60 days, P = 0.0001) when only four babies were discharged on morphine. In contrast, 18 treated period B babies were discharged on morphine. No families were lost to follow up. Compliance with clinic attendance was 92%. CONCLUSIONS: Hospital stay was reduced with establishment of the clinic. The shorter treatment duration before establishment of the clinic could have been related to a lesser abstinence severity or a perceived need of a more rapid weaning prior to discharge. Further studies are needed to assess the impact of this model of care on the health outcome of the narcotic-dependent mother and infant unit.

Caspase-3 activates endo-exonuclease: further evidence for a role of the nuclease in apoptosis.

Single-strand DNase and poly rAase, activities characteristic of endo-exonuclease, were co-activated in nuclear fractions of HL-60 cells by caspase-3. Activation was accompanied by cleavages of large soluble polypeptides (130-185 kDa) and a 65 kDa inactive chromatin-associated polypeptide related to the endo-exonuclease of Neurospora crassa as detected on immunoblots. The major products seen in vitro were a 77 kDa soluble polypeptide and an active chromatin-associated 34 kDa polypeptide. When HL-60 cells were induced to undergo apoptosis by treating with 50 microM etoposide (VP-16) for 4 hours, 77 kDa and 40 kDa polypeptides accumulated in nuclear fractions. Chromatin DNA fragmentation activity was also activated in cytosol and nuclear extract either by pre-treating the cells in vivo with VP-16 or by treating the cytosol in vitro with caspase-3 or dATP and cytochrome c. Endo-exonuclease activated by caspase-3 in cytosol-derived fractions augmented chromatin DNA fragmentation activity in vitro. Endo-exonuclease is proposed to act in vivo in conjunction with the caspase-activated DNase (CAD) to degrade chromatin DNA during apoptosis of HL-60 cells.

Caspase-3-dependent and caspase-3-independent pathways leading to chromatin DNA fragmentation in HL-60 cells.

Apoptosis induced by etoposide (VP-16) in HL-60 cells was confirmed to be caspase-dependent. It was fully inhibited by the broad-spectrum caspase inhibitor Z-VAD-fmk. However, the caspase-3-specific inhibitor Z-DEVDfmk only partially inhibited apoptosis. This indicated that a second caspase is required in vivo for full activation of the apoptotic nucease CAD. Aurin tricarboxylic acid (ATA) did not inhibit VP-16-induced apoptosis. In contrast, apoptosis induced by hydroxychloroquine (HCQ) in HL-60 cells was caspase-3 independent and was fully inhibited by ATA. Thus, CAD does not appear to be involved in chromatin DNA degradation in this case. A second apoptotic nuclease is postulated to degrade the DNA, likely endo- exonuclease, an abundant nuclear enzyme that acts on both DNA and RNA and is present in latent form. HCQ, but not VP-16, stimulated DNA degradation ("laddering") in isolated nuclei. This indicates that the drug can act directly in the nuclei to trigger activation of the second latent apoptotic nuclease.

An investigation of a possible role for mitochondrial nuclease in apoptosis.

Since mitochondrial factors have been implicated in apoptosis, experiments were designed to assess whether or not the potent mitochondrial nuclease could be one of these factors. Nuclei isolated by two different methods were found to contain mitochondrial nuclease in masked form. This nuclease was released by treatment with the non-ionic detergent NP-40 and rendered trypsin-sensitive. It was not removed appreciably from the nuclei by washing and sedimentation of the nuclei through a sucrose cushion. Levels of the mitochondrial nuclease were followed during drug-induced apoptosis. Time courses of apoptosis in cultures of HL-60 cells were monitored by flow cytometry of propidium iodide-stained cells and by agarose gel electrophoresis of extracted DNA. Changes in the inner mitochondrial transmembrane potential were monitored by flow cytometry of chloromethyl-X-Rosamine-stained cells. Apoptosis was induced by treatment with either the chemotherapeutic agent etoposide (VP-16 at 10 microM) over an 8 h period or with the anti-rheumatic agent hydroxychloroquine (HCQ at 0.28 mM) over a 24 h period. These two drugs likely act in different pathways of apoptosis. VP-16 caused loss of the mitochondrial transmembrane potential 1.0-1.5 h before apoptosis was detected. On the other hand, treatment with HCQ caused these processes to occur in parallel possibly indicating that the mitochondrial changes are secondary events. No losses of masked mitochondrial nuclease were detected with either drug treatment during the course of apoptosis. HL-60 mitochondrial DNA was also not degraded during apoptosis induced by either agent. These observations likely explain why the mitochondrial DNA is not degraded and make it unlikely that mitochondrial nuclease plays any role in vivo in chromatin DNA fragmentation.

Induction of apoptosis in peripheral blood lymphocytes following treatment in vitro with hydroxychloroquine.

OBJECTIVE: Defective regulation of apoptosis may be central to the development of autoimmune disorders. This study investigated the possibility that the antirheumatic effect of hydroxycholoroquine (HCQ) may be achieved by up-regulation of apoptosis. METHODS: Peripheral blood lymphocytes collected from normal controls and patients with systemic lupus erythematosus (SLE) were cultured in the presence or absence of a range of concentrations of HCQ. Cells undergoing apoptosis were identified by several standard methods, including morphologic changes, DNA fragmentation, and flow cytometry. For some experiments, lymphocytes were simultaneously stained with antibodies to T cell surface markers and with propidium iodide for dual-stain flow cytometric studies. RESULTS: HCQ was able to induce apoptosis in peripheral blood lymphocytes in a dose- and time-dependent manner. HCQ induced these changes in all T cell subpopulations studied. There was no significant difference between the controls and patients with SLE in terms of the percentage of apoptotic cells detected following treatment with HCQ. CONCLUSION: The present study demonstrated that HCQ induces apoptosis in peripheral blood lymphocytes, which leads to the speculation that HCQ may exert its antirheumatic effect through this mechanism.