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Developmental stress, including low socioeconomic status (SES), can induce dysregulation of the hypothalamic-pituitary-adrenal axis and result in long-term changes in stress reactivity. Children in lower SES conditions often experience more stress than those in other SES groups. There are multiple model systems of early environmental stress (EES), one of which is reduced cage bedding. Here we tested the effects of both prenatal and lactational EES in rats on a range of long-term behavioral and cognitive outcomes. There were persistent reductions in body weight in the EES rats in both sexes. The behavioral results showed no effects on learning and memory using tests of spatial learning or cognitive flexibility in the Morris water maze, egocentric learning in the Cincinnati water maze, or working memory in the radial-arm maze. There were no effects on basic open-field activity, elevated zero-maze, or forced swim test, but EES rats had reduced time in the dark side of the light/dark test. When rats were drug challenged in the open-field with d-amphetamine or MK-801, there were no differential responses to d-amphetamine, but the EES group under responded compared with the drug-induced hyperactivity in the control group in both males and females. The objective was to establish a developmental stress model that induced cognitive deficits and to the extent that this method did not cause such effects it was not the model we sought. However, the data showed several long-term effects of EES, including the reduced response to the irreversible NMDA antagonist MK-801. This effect merits further investigation. • Developmental stress can cause long-term neurobehavioral effects. • Some developmental stressors cause long-term cognitive deficits. • Early Environmental Stress (EES) is reported to cause cognitive deficits in mice. • We tested whether EES in rats causes cognitive and behavioral deficits. • We did not find cognitive deficits in the EES group but found other effects. • The major finding was an under-response to the glutamatergic antagonist MK-801. [ABSTRACT FROM AUTHOR]

In 1979, we tested dietary monosodium glutamate (MSG) for developmental neurotoxicity in rats. The study was recently cited for establishing a No Observable Adverse Effect Level (NOAEL) for MSG as a food additive resulting in a change in the acceptable daily intake (ADI). Therefore, I re-evaluated the study [Vorhees et al.: Toxicol Appl Pharmacol 1979; 50: 267–282]. Sprague-Dawley rats were fed diets containing 0, 1.7, 3.4, or 5.1% MSG prior to conception, throughout gestation and lactation, and the same diets were fed to the offspring until 90 days of age. About 18–20 L were tested per dose with litter and sex factors in data analyses. There were 21 functional tests with 36 dependent variables and 10 body weight and histological outcomes. Of the functional tests, 4 were significant involving 6 effects. Two effects were on swimming ontogeny: one was an improvement and the other an atypical minor delay of no significance. Two effects were on active avoidance: one was a low-dose female-only extinction effect and the other a high-dose male-only acquisition effect, neither providing evidence of consistency. One was on passive avoidance, but was an improvement not a deficit. The last was on open-field rearing in the absence of its normal association with locomotion changes. Thus, it can be concluded, as was done in 1979 and by the U.S. Food and Drug Administration who sponsored the study, that there is no evidence in these data that dietary MSG is developmentally neurotoxic, hence, the study provides no basis for the establishment of a NOAEL and changing the ADI for dietary MSG. [ABSTRACT FROM AUTHOR]

High throughput screens for developmental neurotoxicity (DN) will facilitate evaluation of chemicals and can be used to prioritize those designated for follow-up. DN is evaluated under different guidelines. Those for drugs generally include peri- and postnatal studies and juvenile toxicity studies. For pesticides and commercial chemicals, when triggered, include developmental neurotoxicity studies (DNT) and extended one-generation reproductive toxicity studies. Raffaele et al. (2010) reviewed 69 pesticide DNT studies and found two of the four behavioral tests underperformed. There are now many epidemiological studies on children showing adverse neurocognitive effects, yet guideline DN studies fail to assess most of the functions affected in children; nor do DN guidelines reflect the advances in brain structure-function relationships from neuroscience. By reducing the number of test ages, removing underperforming tests and replacing them with tests that assess cognitive abilities relevant to children, the value of DN protocols can be improved. Testing for the brain networks that mediate higher cognitive functions need to include assessments of working memory, attention, long-term memory (explicit, implicit, and emotional), and executive functions such as cognitive flexibility. The current DNT focus on what can be measured should be replaced with what should be measured. With the wealth of data available from human studies and neuroscience, the recommendation is made for changes to make DN studies better focused on human-relevant functions using tests of proven validity that assess comparable functions to tests used in children. Such changes will provide regulatory authorities with more relevant data. [ABSTRACT FROM AUTHOR]

Advantageous maneuvering through the environment to find food and avoid or escape danger is central to survival of most animal species. The ability to do so depends on learning and remembering different locations, especially home-base. This capacity is encoded in the brain by two systems: one using cues outside the organism (distal cues), allocentric navigation, and one using self-movement, internal cues (proximal cues), for egocentric navigation. Whereas allocentric navigation involves the hippocampus, entorhinal cortex, and surrounding structures, egocentric navigation involves the dorsal striatum and connected structures; in humans this system encodes routes and integrated paths and when over-learned, becomes procedural memory. Allocentric assessment methods have been extensively reviewed elsewhere. The purpose of this paper is to review one specific method for assessing egocentric, route-based navigation in rats: the Cincinnati water maze (CWM). The test is an asymmetric multiple-T maze arranged in such a way that rats must learn to find path openings along walls rather at ends in order to reach the goal. Failing to do this leads to cul-de-sacs and repeated errors. The task may be learned in the light or dark, but in the dark, wherein distal cues are eliminated, provides the best assessment of egocentric navigation. When used in conjunction with tests of other types of learning, such as allocentric navigation, the CWM provides a balanced approach to assessing the two major forms of navigational learning and memory found in mammals. [ABSTRACT FROM AUTHOR]

Cognitive tests of learning and memory (L&M) have been required by U.S. Environmental Protection Agency (EPA) developmental neurotoxicity test (DNT) guidelines for more than two decades. To evaluate the utility of these guidelines, the EPA reviewed 69 pesticide DNT studies. This review found that the DNT provided or could provide the point-of-departure for risk assessment by showing the Lowest Observable Adverse Effect Level (LOAEL) in 28 of these studies in relation to other reported end points. Among the behavioral tests, locomotor activity and auditory/acoustic startle provided the most LOAELs, and tests of cognitive function and the Functional Observational Battery (FOB) the fewest. Two issues arose from the review: (1) what is the relative utility of cognitive tests versus tests of unconditioned behavior, and (2) how might cognitive tests be improved? The EPA sponsored a symposium to address this. Bushnell reviewed studies in which both screening (locomotor activity, FOB, reflex ontogeny, etc.) and complex tests (those requiring training) were used within the same study; he found relatively little evidence that complex tests provided a LOAEL lower than screening tests (with exceptions). Levin reviewed reasons for including cognitive tests in regulatory studies and methods and evidence for the radial arm maze and its place in developmental neurotoxicity assessments. Driscoll and Strupp reviewed the value of serial reaction time operant methods for assessing executive function in developmental neurotoxicity studies. Vorhees and Williams reviewed the value of allocentric (spatial) and egocentric cognitive tests and presented methods for using the Morris water maze for spatial and the Cincinnati water maze for egocentric cognitive assessment. They also reviewed the possible use of water radial mazes. The relatively lower impact of cognitive tests in previous DNT studies in the face of the frequency of human complaints of chemical-induced cognitive dysfunction indicates that animal cognitive tests need improvement. The contributors to this symposium suggest that if the guidelines are updated, they be made more specific by recommending preferred tests and providing greater detail on key characteristics of such tests. Additionally, it is recommended that guidance be developed to address important issues with cognitive tests and to provide the information needed to improve the design, conduct, and interpretation of tests of higher function within a regulatory context. These steps will maximize the value of cognitive tests for use in hazard evaluation and risk assessment. [ABSTRACT FROM AUTHOR]

There are a variety of chemicals, including pharmaceuticals, that alter neurobehavior following developmental exposure and guidelines for the conduct of studies to detect such effects by statute in the United States and Europe. Guidelines for Developmental Neurotoxicity Testing (DNT) studies issued by the U.S. Environmental Protection Agency (EPA) under prevailing law and European Organization for Economic Cooperation and Development (OECD) recommendations to member countries provide that such studies include a series of neurobehavioral and neuropathological assessments. Among these are assessment of cognitive function, specifically learning and memory. After reviewing 69 DNT studies submitted to the EPA, tests of learning and memory were noted to have detected the lowest observed adverse effect level (LOAELs) less frequently than behavioral tests of locomotor activity and acoustic/auditory startle, but slightly more than for the developmental Functional Observational Battery (devFOB; which is less extensive than the full FOB), but the reasons for the lower LOAEL detection rate for learning and memory assessment could not be determined. A major concern identified in the review, however, was the adequacy of the methods employed in these studies rather than on the importance of learning and memory to the proper assessment of brain function. Accordingly, a symposium was conducted to consider how the guidelines for tests of learning and memory might be improved. Four laboratories with established histories investigating the effects of chemical exposures during development on learning, memory, and attention, were invited to review the topic and offer recommendations, both theoretical and practical, on approaches to improve the assessment of these vital CNS functions. Reviewers were asked to recommend methods that are grounded in functional importance to CNS integrity, well-validated, reliable, and amenable to the context of regulatory studies as well as to basic research on the underlying processes they measure. This Introduction sets the stage for the reviews by providing the background and regulatory context for improved tests for learning and memory in DNT and other regulatory studies, such as single- or multi-generational studies where similar methods are incorporated. [ABSTRACT FROM AUTHOR]

Maneuvering safely through the environment is central to survival of all animals. The ability to do this depends on learning and remembering locations. This capacity is encoded in the brain by two systems: one using cues outside the organism (distal cues), allocentric navigation, and one using self-movement, internal cues and sometimes proximal cues, egocentric navigation. Allocentric navigation involves the hippocampus, entorhinal cortex, and surrounding structures (e.g., subiculum); in humans this system encodes declarative memory (allocentric, semantic, and episodic, i.e., memory for people, places, things, and events). This form of memory is assessed in laboratory animals by many methods, but predominantly the Morris water maze (MWM). Egocentric navigation involves the dorsal striatum and connected structures; in humans this system encodes routes and integrated paths and when over-learned becomes implicit or procedural memory. Several allocentric methods for rodents are reviewed and compared with the MWM with particular focus on the Cincinnati water maze (CWM). MWM advantages include minimal training, no food deprivation, ease of testing, reliable learning, insensitivity to differences in body weight and appetite, absence of non-performers, control methods for performance effects, repeated testing capability and other factors that make this test well-suited for regulatory studies. MWM limitations are also reviewed. Evidence-based MWM design and testing methods are presented. On balance, the MWM is arguably the preferred test for assessing learning and memory in basic research and regulatory studies and the CWM is recommended if two tests can be accommodated so that both allocentric (MWM) and egocentric (CWM) learning and memory can be effectively and efficiently assessed. [ABSTRACT FROM AUTHOR]

Prenatal maternal immune activation increases risk for schizophrenia and/or autism. Previous data suggest that maternal weight change in response to the immune activator polyinosinic–polycytidylic (Poly IC) in rats influences the severity of effect in the offspring as does the exposure period. We treated gravid Sprague–Dawley rats from E14 to 18 with 8 mg/kg/day Poly IC or saline. The Poly IC group was divided into those that gained the least weight or lost (Poly IC (L)) and those that gained the most (Poly IC (H)) weight. There were no effects of Poly IC on anxiety (elevated zero-maze, open-field, object burying), or Morris water maze cued learning or working memory or Cincinnati water maze egocentric learning. The Poly IC (H) group males had decreased acoustic startle whereas Poly IC (L) females had reduced startle and increased PPI. Poly IC offspring showed exaggerated hyperactivity in response to amphetamine (primarily in the Poly IC (H) group) and attenuated hyperactivity in response to MK-801 challenge (primarily in the Poly IC (L) group). Poly IC (L) males showed reduced cued conditioned freezing; both sexes showed less time in the dark in a light–dark test, and the Poly IC groups showed impaired Morris water maze hidden platform acquisition and probe performance. The data demonstrate that offspring from the most affected dams were more affected than those from less reactive dams indicating that degree of maternal immune activation predicts severity of effects on offspring behavior. [ABSTRACT FROM AUTHOR]

This commentary is in response to the remarks of Drs. Christina Sobin, Mari Golub, and David Herr on the Special Issue of this Journal entitled “Assessment of Learning, Memory, and Attention in Developmental Neurotoxicology Regulatory Testing.” I endorse the views expressed by Drs. Sobin, Golub, and Herr and add some discussion on a recent Organization for Economic Cooperation and Development (OECD) guideline, the Extended One Generation Reproductive Toxicology Guideline (OECD 446), in which testing for higher cognitive function (learning and memory) has been eliminated. The case against this decision is offered. It is noted that deficits in higher cognitive function are one of the hallmarks of human studies that find neurobehavioral toxicity in children after exposure to environmental agents such as lead, methylmercury, PCB, pesticides, and other environmental agents. It is noted that the OECD decision is at variance with the views of the scientific community in this field, including those of Drs. Sobin, Golub, and Herr. Why OECD took such action without the advice and consent of the field of developmental neurotoxicology is deeply concerning and potentially hazardous to children. I also endorse Dr. Herr's recommendation that in the future the Environmental Protection Agency negotiate study designs in advance with submitters as the Food and Drug Administration does to improve data quality for all neurobehavioral methods, and especially for tests of learning and memory that have not been adequately conducted in many past studies. [ABSTRACT FROM AUTHOR]

Maneuvering safely through the environment is central to survival of all animals. The ability to do this depends on learning and remembering locations. This capacity is encoded in the brain by two systems: one using cues outside the organism (distal cues), allocentric navigation, and one using self-movement, internal cues and sometimes proximal cues, egocentric navigation. Allocentric navigation involves the hippocampus, entorhinal cortex, and surrounding structures (e.g., subiculum); in humans this system encodes declarative memory (allocentric, semantic, and episodic, i.e., memory for people, places, things, and events). This form of memory is assessed in laboratory animals by many methods, but predominantly the Morris water maze (MWM). Egocentric navigation involves the dorsal striatum and connected structures; in humans this system encodes routes and integrated paths and when over-learned becomes implicit or procedural memory. Several allocentric methods for rodents are reviewed and compared with the MWM with particular focus on the Cincinnati water maze (CWM). MWM advantages include minimal training, no food deprivation, ease of testing, reliable learning, insensitivity to differences in body weight and appetite, absence of non-performers, control methods for performance effects, repeated testing capability and other factors that make this test well-suited for regulatory studies. MWM limitations are also reviewed. Evidence-based MWM design and testing methods are presented. On balance, the MWM is arguably the preferred test for assessing learning and memory in basic research and regulatory studies and the CWM is recommended if two tests can be accommodated so that both allocentric (MWM) and egocentric (CWM) learning and memory can be effectively and efficiently assessed. [ABSTRACT FROM AUTHOR]

Prenatal maternal immune activation has been used to test the neurodevelopmental hypothesis of schizophrenia. Most of the data are in mouse models; far less is available for rats. We previously showed that maternal weight change in response to the immune activator polyinosinic-polycytidylic acid (Poly IC) in rats differentially affects offspring. Therefore, we treated gravid Harlan Sprague-Dawley rats i.p. on embryonic day 14 with 8 mg/kg of Poly IC or Saline. The Poly IC group was divided into those that lost or gained the least weight, Poly IC (L), versus those that gained the most weight, Poly IC (H), following treatment. The study design controlled for litter size, litter sampling, sex distribution, and test experience. We found no effects of Poly IC on elevated zero maze, open-field activity, object burying, light-dark test, straight channel swimming, Morris water maze spatial acquisition, reversal, or shift navigation or spatial working or reference memory, or conditioned contextual or cued fear or latent inhibition. The Poly IC (H) group showed a significant decrease in the rate of route-based learning when visible cues were unavailable in the Cincinnati water maze and reduced prepulse inhibition of acoustic startle in females, but not males. The Poly IC (L) group exhibited altered responses to acute pharmacological challenges: exaggerated hyperactivity in response to (+)-amphetamine and an attenuated hyperactivity in response to MK-801. This model did not exhibit the cognitive, or latent inhibition deficits reported in Poly IC-treated rats but showed changes in response to drugs acting on neurotransmitter systems implicated in the pathophysiology of schizophrenia (dopaminergic hyperfunction and glutamatergic hypofunction). Synapse 2012. © 2012 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

The article discusses the experiment conducted that compared ohenylethylamines such as (+)-methamphetamine (MA), (±)-3,4-methylenedioxymethamphetamine (MDMA), (+)-amphetamine (AMPH), and (±)-fenfluramine (FEN) on Cincinnati water maze (CWM) performance of rats. AMPH and MA were shown to have induced an increase in CWM errors and latency in achieving the goal, with no difference in swim speed, while learning was not significantly altered by MDMA and FEN.

Polychlorinated biphenyls (PCBs) are persistent toxic pollutants occurring as complex mixtures in the environment. Humans are known genetically to have > 60-fold differences in hepatic cytochrome P450 1A2 (CYP1A2) levels and > 12-fold differences in aryl hydrocarbon receptor (AHR) affinity, both of which could affect PCB pharmacokinetics. Thus, we compared Ahrb1_Cyp1a2(+/+) high-affinity AHR wild-type, Ahrd_Cyp1a2(+/+) poor affinity AHR wild-type, Ahrb1_Cyp1a2(−/−) knockout, and Ahrd_Cyp1a2(−/−) knockout mouse lines. We chose a mixture of three coplanar and five noncoplanar PCBs to reproduce that seen in human tissues, breast milk, and the food supply. The mixture was given by gavage to the mother on gestational day 10.5 (GD10.5) and postnatal day 5 (PND5); tissues were collected from pups and mothers at GD11.5, GD18.5, PND6, PND13, and PND28. Ahrb1_Cyp1a2(−/−) pups showed lower weight at birth and slower rate of growth postnatally. Absence of CYP1A2 resulted in significant splenic atrophy at PND13 and PND28. Presence of high-affinity AHR enhanced thymic atrophy and liver hypertrophy in the pups. Concentrations of each congener were analyzed at all time points: maximal noncoplanar congener levels in maternal tissues were observed from GD18 until PND6, whereas the highest levels in pups were found between PND6 and PND28. Coplanar PCB concentrations were generally higher in Ahrd-containing pup tissues; these findings are consistent with earlier studies demonstrating the crucial importance of AHR-mediated inducible CYP1 in the gastrointestinal tract as a means of detoxication of oral planar polycyclic aromatic hydrocarbons. [ABSTRACT FROM AUTHOR]

Abstract: Rats treated with (+)-methamphetamine (MA) on postnatal days (P) 11–20 exhibit long-term spatial and path integration (Morris water maze (MWM) and Cincinnati water maze (CWM)) learning deficits whereas those treated on P1–10 do not. MA treatment increases corticosterone release in an age-dependent U-shaped pattern that corresponds to the stress hyporesponsive period (SHRP; P4–15). Here we tested the hypothesis that the cognitive effects induced by MA are associated with treatment that begins within the SHRP. Three treatment regimens were compared, P1–10, P6–15, and P11–20. One male/female pair/litter received 0, 10, or 25mg/kg MA/dose (four doses/day at 2h intervals given s.c. with 19–21 litters/regimen). Locomotor activity and acoustic startle were tested as behaviors not predicted to be associated with the SHRP. Cincinnati and Morris water maze findings were consistent with the hypothesis in that MA-treated animals exposed from P6–15 or P11–20 showed impaired learning compared to those exposed from P1–10; however, on probe trials in the Morris water maze, MA-induced memory impairments were not regimen-specific and were contributed to by all treatment regimens. All MA treatment regimens induced reductions in locomotor activity and acoustic startle facilitation as expected. No differential effect on prepulse trials was seen suggesting no impairment in sensory gating. Cognitive deficits from neonatal MA treatment are associated with the SHRP and may be the product of hypothalamic–pituitary–adrenal (HPA) axis dysregulation during critical periods of brain development. [Copyright &y& Elsevier]

Abstract: Developmental exposure to the dopamine D2/3 receptor agonist quinpirole is reported to induce D2 priming, impair Morris water maze performance, reduce acoustic startle prepulse inhibition (PPI), and alter locomotor activity. We treated rats from postnatal days 1–21 with the dose reported to induce these effects, 1.0 mg/kg/day, and two higher doses, 2.0 and 4.0 mg/kg/day, or saline. Offspring were tested in the Morris water maze, PPI, exploratory locomotor activity, activity after quinpirole and (+)-methamphetamine challenge, elevated zero maze, light-dark box, marble burying, straight channel swimming, and Cincinnati water maze. In the Morris water maze, all quinpirole groups had longer latencies on test days 3–5 of acquisition, but no effects on reversal or shifted-reduced platform trials. The quinpirole 4.0 mg/kg group had significantly reduced mean search distances on probe trials when combined across the 3 phases of testing but not separately. The male 4.0 mg/kg quinpirole group showed a greater increase in methamphetamine-stimulated activity during the first 10 min after drug challenge but not in the remainder of the 2 h test. No quinpirole effects were found for light-dark box, marble burying, exploratory locomotor activity, straight channel, Cincinnati water maze, or locomotor activity after quinpirole challenge. No effects were found on most measures in the elevated zero maze however the quinpirole 4.0 mg/kg females had longer latencies to enter an open quadrant. The results partially support prior Morris maze deficits induced by developmental quinpirole treatment but little evidence of dopamine D2/3 priming was found using locomotor activity with quinpirole or methamphetamine challenge or acoustic startle/PPI. The limited comparability to published data using developmental quinpirole exposure may be attributable to differences in experimental procedures or may be the result of quinpirole having limited effects. The data suggest that caution is warranted concerning the developmental efficacy of quinpirole. [Copyright &y& Elsevier]

Abstract: Postnatal day (P)11–20 (+)-methamphetamine (MA) treatment impairs spatial learning and reference memory in the Morris water maze, but has marginal effects on learning in a labyrinthine maze. A subsequent experiment showed that MA treatment on P11–15, but not P16–20, is sufficient to induce Morris maze deficits. Here we tested the effects of P11–15 MA treatment under two different rearing conditions on Morris maze performance and path integration learning in the Cincinnati water maze in which distal cues were unavailable by using infrared illumination. Littermates were treated with 0, 10, 15, 20, or 25mg/kg MA×4/day (2h intervals). Half the litters were reared under standard housing conditions and half under partial enrichment by adding stainless steel enclosures. All MA groups showed impaired Cincinnati water maze performance with no significant effects of rearing condition. In the Morris maze, the MA-25 group showed impaired spatial acquisition, reversal, and small platform learning. Enrichment significantly improved Morris maze acquisition in all groups but did not interact with treatment. The male MA-25 group was also impaired on probe trial performance after acquisition and on small platform trials. A narrow window of MA treatment (P11–15) induces impaired path integration learning irrespective of dose within the range tested but impairments in spatial learning are dependent on dose. The results demonstrate that a narrower exposure window (5 days) changes the long-term effects of MA treatment compared to longer exposures (10 days). [Copyright &y& Elsevier]

We previously demonstrated that postnatal day 11-20 ±3,4-methylenedioxymethamphetamine (MDMA) exposure reduces locomotor activity and impairs path integration and spatial learning independent of the effects on activity. The effects were seen when the drug was administered twice per day, but the optimal dosing regimen is unknown. We tested whether the same total daily dose of MDMA administered in different patterns would equally affect later behavior. A split-litter design (15 litters) was used with one male/female pair per litter receiving one of four treatment regimens. All offspring received four injections per day on P11-20 as follows: 40 × 1 (40 mg/kg MDMA × 1 + saline × 3), 20 × 2 (20 mg/kg MDMA × 2 + saline × 2), 10 × 4 (10 mg/kg MDMA × 4), or Saline (saline × 4). Doses were spaced 2 h apart. Group 40 × 1 received MDMA as the first daily dose followed by three saline doses; group 20 × 2 received MDMA as the first and last dose and saline for the middle two doses; group 10 × 4 received MDMA for all four doses; and the saline group received saline for all four doses. Regardless of dose schedule, all groups treated with MDMA exhibited reduced locomotor activity. No MDMA effects were found on swimming ability in a straight channel. Modest MDMA effects were found on Barnes maze performance. The major findings were that the 40 × 1 and 20 × 2 MDMA groups showed impaired Cincinnati multiple T-water-maze learning and the 10 × 4 and 20 × 2 MDMA groups showed impaired Morris water maze learning. The results suggest that MDMA dose distribution has a long-term differential effect on different types of learning. Dose distribution warrants greater attention in the design of developmental drug studies along with the standard considerations of dose and age. Synapse 61:488-499, 2007. © 2007 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Highlights the importance of toxicity testing of pesticides and industrial chemicals. Consequences of exposure to lead, methyl mercury, polychlorinated biphenyls and tobacco; Origin and evolution of developmental neurotoxicity (DNT) tests; Actions taken by the U.S. government to protect children from environmental toxins.

Abstract: We have previously shown that P11–20 treatment with d-methamphetamine (MA) induces impaired spatial navigation in the Morris water maze (MWM), whereas P1–10 treatment does not. Little is known about the long-term behavioral consequences of MA during juvenile, adolescent, and early adult brain development. In dose–response experiments, we tested successive 10-day intervals of exposure to MA in rats (P21–30, P31–40, P41–50, and P51–60; four doses per day). MA dosing prior to P21 produces little or no toxicity; however, we observed an increased toxicity with advancing age. Across-age comparisons revealed no MWM acquisition or Cincinnati water maze (CWM) effects after MA treatment on P21–30 (2.5–10 mg/kg/dose), P31–40 (1.25–7.5 mg/kg/dose), or P51–60 (1.25–5.0 mg/kg/dose); however, significantly impaired MWM acquisition was observed after P41–50 MA treatment at the highest dose (6.25 mg/kg/dose). Learning in the CWM was also impaired in this group. No effects were seen at 1.25, 2.5, or 5 mg/kg/dose following P41–50 MA treatment. MWM reversal learning trials after P41–50 treatment showed a trend towards longer latency in all MA dose groups, but no effect on double-reversal trials. Reversal and double-reversal also showed no effects at the other exposure ages. No differences in straight channel swimming or cued learning in the MWM were seen after MA treatment at any exposure age. P41–50 is the periadolescent stage of brain development in rodents. The effects observed at this age may suggest a previously unrecognized period of susceptibility for MA-induced cognitive deficits. [Copyright &y& Elsevier]

3,4-Methylenedioxymethamphetamine (MDMA) in previous experiments has been shown to induce long-term spatial and sequential learning and memory deficits in adult offspring after exposure to the drug on postnatal (P) days 11–20, but not after exposure on P1–10. Herein we further tested for the effects of MDMA (0, 5, 10 or 20mg/kg × 2/day) after exposure on P11–20 on reference and working memory in the Morris water maze (MWM), on reference memory in the Barnes maze, and on cued learning in the visible platform version of the MWM. The MWM and Barnes mazes were counterbalanced such that half the litters received the MWM-first and the other half received the Barnes maze first. Effects on MWM performance as a function of test order were observed. For animals that received the Barnes maze first, spatial MWM learning and memory trends were seen but they were not significantly different between MDMA groups and saline controls. For those receiving the MWM-first, there are consistent impairments on all measures in the MDMA groups compared to controls on MWM performance (latency, path length, and cumulative distance from the goal). On probe trials, MDMA animals receiving the MWM-first showed increased distance from the target site compared to controls. There were no MDMA effects seen on cued trials in the MWM or on straight channel swimming trials regardless of test order, indicating that MDMA had no effects on swimming ability or on the skills needed to learn the MWM. Similarly, there were no effects of MDMA on MWM working memory regardless of test order. No MDMA effects on the Barnes maze were found regardless of test order, however, the interpretation of this finding was compromised by the poor performance of the animals on this task. [Copyright &y& Elsevier]

Spatial learning and memory deficits in a water maze have been observed in adult animals exposed to a regimen of 4 daily doses of d-methamphetamine (MA) at 2 h intervals from postnatal day 11 to 20. An interpretational issue for these long-term effects of MA is whether they are truly spatial deficits or are secondary to alterations in sensorimotor systems. In this experiment, we evaluated the effects of a pretraining procedure shown to minimize the influence of drug-induced sensorimotor deficits. Animals within a litter were treated with MA or saline. Animals were either pretrained for nonspatial task requirements in the water maze (i.e., swimming and platform climbing) or were naı¨ve to the task. Animals that received the pretraining did better than the naı¨ve animals. The naı¨ve MA animals performed worse than the naı¨ve control animals as previously observed. By contrast, no difference in search time was noted between pretrained MA- and SAL-treated animals during the acquisition phase of testing. When the platform was relocated in a novel position, spatial learning was impaired for MA animals, regardless of pretraining. No increase in the number of platform nonrecognition events (swimovers, deflections, or jump-offs) occurred among pretrained or naı¨ve groups compared to controls. These data suggest that sensorimotor deficits do not account for the spatial learning and memory deficits in animals exposed neonatally to MA. [Copyright &y& Elsevier]

Apoptosis is an important process in the development and function of the central nervous system (CNS). To study the role of DNA fragmentation factor 45 (DFF45/ICAD) in CNS function, we previously generated DFF45 knockout mice. We found that whereas they exhibit apparently normal CNS development, DFF45 knockout mice exhibit an increased number of granule cells in the dentate gyrus and enhanced spatial learning and memory compared to wild-type mice in a Morris water maze test. In this study, we examined the performance of the DFF45 knockout mice in a novel object recognition task to measure short-term nonspatial memory that is believed to depend on the hippocampal formation. Both wild-type and DFF45 knockout mice exhibited novel object recognition 1 h posttraining. However, whereas wild-type mice no longer did so, DFF45 knockout mice were still able to differentiate the novel versus the familiar object 3 h posttraining. The longer memory retention in DFF45 knockout mice did not last up to 24 h as neither wild-type nor DFF45 knockout mice demonstrated novel object recognition 24 h posttraining. These results suggest that a lack of DFF45 facilitates hippocampus-dependent nonspatial memory, as well as hippocampus-dependent spatial memory. [Copyright &y& Elsevier]

Developmental neurotoxicity (DNT) studies could benefit from revisions to study design, data analysis, and some behavioral test methods to enhance reproducibility. The Environmental Protection Agency (EPA) reviewed 69 studies submitted to the Office of Pesticide Programs. Two of the behavioral tests identified the lowest observable adverse effect level (LOAEL) 20 and 13 times, respectively, while the other two tests identified the LOAEL only 3 and 4 times, respectively. The EPA review showed that the functional observational battery (FOB) was least effective at detecting the LOAEL, whereas tests of learning and memory (L&M) had methodological shortcomings. Human neurodevelopmental toxicity studies over the past 30 years show that most of the adverse effects are on higher cognitive functions such as L&M. The results of human studies together with structure-function relationships from neuroscience, suggest that tests of working memory, spatial navigation/memory, and egocentric navigation/memory should be added to guideline studies. Collectively, the above suggest that EPA and EU DNT studies would better reflect human findings and be more relevant to children by aligning L&M tests to the same domains that are affected in children, removing less useful methods (FOB), and using newer statistical models to better account for random factors of litter and litter × sex. Common issues in study design and data analyses are discussed: sample size, random group assignment, blinding, elimination of subjective rating methods, avoiding confirmation bias, more complete reporting of species, housing, test protocols, age, test order, and litter effects. Litter in DNT studies should at least be included as a random factor in ANOVA models and may benefit from inclusion of litter × sex as random factors. • Developmental neurotoxicity (DNT) studies need to improve rigor and reproducibility. • Regulatory DNT study guidelines need revision based on data in children. • Small N, lack of litter control, and correlated variables compromise DNT studies. • Litter and litter x sex should be random factors in ANOVA models. • Randomization, blinding, confirmation bias, and "helping" animals remain issues. [ABSTRACT FROM AUTHOR]

Neurotoxicity test batteries are collections of individual tests assembled for the purpose of detecting multiple aspects of CNS dysfunction in experimental animals. The use of such test batteries has grown substantially over the last 20 years and much has been learned from this experience about the design, analysis, and interpretation of test batteries. This review discusses the advantages and disadvantages of test batteries and the design consideration that enter into the process of developing a battery. At the heart of test battery development is a series of balancing factors that must be weighed by the investigator in order to determine what types of tests to include in a given test battery. While test batteries offer some distinct advantages, their limitations are often overlooked in the early stages of planning. The goal of this review is to present these limiting factors so that investigators may understand the problems they will confront both before and after the experiment is performed in order to help maximize the chances of a successful outcome. [ABSTRACT FROM AUTHOR]

Two developmental stressors were compared in preweaning rats exposed to either one stressor or both. Stressors were barren cage rearing or maternal separation (pup isolation). 40 gravid Sprague-Dawley CD/IGS rats were randomly assigned to two cage conditions: standard (Std) cage or barren cage (Bar), 20 litters/condition throughout gestation and lactation. After delivery, litters were randomly culled to 4 males and 4 females. The second stressor was maternal separation: Two male/female pairs per litter were isolated from their dam 4 h/day (Iso) and two pairs were not (Norm). Hence, there were 4 conditions: Std-Norm, Std-Iso, Bar-Norm, and Bar-Iso. One pair/litter/stress condition received the following: elevated zero-maze (EZM), open-field, swim channel, Cincinnati water maze, conditioned fear, and open-field with methamphetamine challenge. The second pair/litter/condition received the light-dark test, swim channel, Morris water maze, forced swim, and EZM with diazepam challenge. Barren rearing reduced EZM time-in-open, whereas isolation rearing reduced open-field activity in males and increased it in females. Effects on straight channel swimming were minor. In the Cincinnati water maze test of egocentric learning, isolation rearing increased errors whereas barren cage housing reduced errors in combination with normal rearing. Barren cage with maternal separation (pup isolation) increased Cincinnati water maze escape latency but not errors. Barren cage housing reduced hyperactivity in response to methamphetamine. Isolation rearing increased time in open in the EZM after diazepam challenge. Trends were seen in the Morris water maze. These suggested that barren cage and isolation rearing in combination reduced latency on acquisition on days 1 and 2 in males, whereas females had increased latency on days 2 and 3. Combined exposure to two developmental stressors did not induce additive or synergistic effects, however the data show that these stressors had long-term effects with some evidence that the combination of both caused effects when either stressor alone did not, but synergism was not observed. • Compare barren cage rearing to maternal separation (pup isolation) on behavior. • Barren cage rearing increased anxiety-like behavior in the elevated zero maze. • Isolation reduced open-field activity in males and increased it in females. • Isolation increased Cincinnati water maze (CWM) errors, barren cage reduced them. • Barren + isolation increased anxiety-like behavior and decreased CWM performance. [ABSTRACT FROM AUTHOR]

The importance of litter effects (clustering of variance among offspring in rodents) has been known for decades. The standard approach was to treat the entire litter as a unit or to select one male and one female from each litter to prevent oversampling. These methods work but are imperfect. Treating the litter as a whole fails to use valuable interindividual differences among offspring, and selecting representative pups fails to use all the data available. Golub and Sobin [ https://doi.org/10.1016/j.ntt.2019.106841 ] address this using a better method. They show that using litter as a random factor in mixed linear models resolves this conundrum. As they demonstrate, such models control for litter clustering by partitioning litter variance from error variance. This reduces error variance and increases the power of F-tests of the independent variable(s). In our experience, this is the optimal solution. But as good as mixed linear models are when used with litter as a random factor, if other aspects of the experimental design are not appropriate, this cannot compensate for threats to validity from small sample sizes, dams not strictly randomly assigned to groups, repeated measure covariance structures not appropriately modeled, interactions not properly sliced, or a posteriori group comparisons not controlled for multiple comparisons. Appropriate handling of litter is only one consideration of experimental design and statistical analysis that when used in combination lead to valid, reproducible data. • In prenatal and many preweaning studies dam/litter is the unit of analysis. • It is known that within litter variance is less than between litter variance. • Golub and Sobin show the optimal statistical method to account for litter effects. • Litter should be a random factor in mixed linear ANOVA models. • Failure to control litter effects is a serious flaw found in many studies. [ABSTRACT FROM AUTHOR]

• ADHD is polygenic with multiple small effect gene variants contributing. • ADHD is associated with some environmental chemical or drug exposures. • Selectively bred and gene knockout models are reviewed. • Genetic and environmental models have been studied separately. • No two or three combination knockout/knockdown models have been described. Attention deficit hyperactivity disorder (ADHD) is a polygenic neurodevelopmental disorder that affects 8–12 % of children and >4 % of adults. Environmental factors are believed to interact with genetic predispositions to increase susceptibility to ADHD. No existing rodent model captures all aspects of ADHD, but several show promise. The main genetic models are the spontaneous hypertensive rat, dopamine transporter knock-out (KO) mice, dopamine receptor subtype KO mice, Snap-25 KO mice, guanylyl cyclase-c KO mice, and latrophilin-3 KO mice and rats. Environmental factors thought to contribute to ADHD include ethanol, nicotine, PCBs, lead (Pb), ionizing irradiation, 6-hydroxydopamine, neonatal hypoxia, some pesticides, and organic pollutants. Model validation criteria are outlined, and current genetic models evaluated against these criteria. Future research should explore induced multiple gene KOs given that ADHD is polygenic and epigenetic contributions. Furthermore, genetic models should be combined with environmental agents to test for interactions. [ABSTRACT FROM AUTHOR]

• Latrophilins are adhesion G-protein-coupled receptors. • Latrophilin-3 (Lphn3) is a trans-synaptic regulator of synaptic function. • 21 gene variants of LPHN3 are linked to ADHD. • Lphn3 disruption in Drosophila , zebrafish, mice, and rats result in hyperactivity. • Lphn3 KO rats show selective cognitive deficits and dopamine abnormalities. Latrophilin-3 (LPHN3), a G-protein-coupled receptor belonging to the adhesion subfamily, is a regulator of synaptic function and maintenance in brain regions that mediate locomotor activity, attention, and memory for location and path. Variants of LPHN3 are associated with increased risk for attention deficit hyperactivity disorder (ADHD) in some patients. Here we review the role of LPHN3 in the central nervous system (CNS). We describe synaptic localization of LPHN3, its trans-synaptic binding partners, links to neurodevelopmental disorders, animal models of Lphn3 disruption in different species, and evidence that LPHN3 is involved in cognition as well as activity and attention. The evidence shows that LPHN3 plays a more significant role in neuroplasticity than previously appreciated. [ABSTRACT FROM AUTHOR]

Children exposed to methamphetamine (MA) in utero have cognitive deficits. MA administration in rats for 5–10 days between postnatal days (P)6 and 20 produces cognitive deficits. The purpose of this study was to determine if extending MA administration by 5 days within P6–20 would exacerbate allocentric (Morris water maze) and egocentric (Cincinnati water maze) learning deficits. Sprague Dawley female and male offspring (split‐litter design) were administered saline (SAL) or MA (10 mg/kg) four times daily from P6 to 20 to create four groups: (a) SAL from P6 to 20, (b) MA from P6 to 20 (MA6–20), (c) MA from P6 to 15 (MA6–15), or (d) MA from P11 to 20 (MA11–20); the latter groups received saline on days they did not receive MA. Egocentric, allocentric, and conditioned freezing tests began on P60. The MA6–15 and MA6–20 groups showed egocentric deficits, all MA groups had allocentric deficits but no differences in conditioned freezing compared with SAL controls. The MA6–15 and MA6–20 groups had similar deficits in learning and memory that were larger than in the MA11–20 group. Learning in both mazes was sex dependent, but no interactions with MA were found. The data demonstrate that extending the exposure period of MA beyond the sensitive periods (P6–15 and P11–20) did not exacerbate the cognitive deficits. The purpose of this study was to determine if extending MA administration by 50% of 10 days within the critical period of postnatal days (P)6–20 would exacerbate allocentric and egocentric learning deficits. Similar egocentric deficits were found when dosing started on P6 and all MA groups had allocentric deficits compared with SAL controls. The data demonstrate that extending the exposure period of MA during the sensitive periods (P6–15 and P11–20) did not exacerbate the cognitive deficits. [ABSTRACT FROM AUTHOR]

Deltamethrin (DLM) is a commonly used pesticide that helps to control crop destruction, disease, and nuisance insects. In rodents DLM can produce choreoathetosis, salivation, and decreased acoustic startle responses (ASR). Herein, adult Sprague Dawley rats were assessed for ASR 2 h after DLM delivered in 5 ml/kg corn oil, however no decrease was observed. Therefore, a test-retest protocol was used to reduce variability, and the effects on ASR on postnatal day 15 (P15) and adult rats were assessed 2, 4, 6, and 8 h after DLM administration (0, 1, 2, or 4 mg/kg for P15 rats and 0, 2, 8, or 25 mg/kg for adults). In a separate set of rats identically treated, DLM levels were determined in blood and brain. DLM (8 or 25 mg/kg) in adult rats decreased ASR up to 4 h, whereas in P15 rats decreases were observed between 2 and 8 h. The adult 25 mg/kg group showed consistent signs of salivation and tremor, whereas in P15 rats salivation was observed in the 2 and 4 mg/kg groups and tremor was observed at all doses over the 8-h period. Mortality was observed in all P15 dose groups but not in adults. Dose-dependent increases of DLM in blood and brain regardless of age were observed. At approximately equivalent whole brain concentrations, effects were more pronounced in P15 rats than in adult rats. Comparable brain levels of DLM do not explain differences in ASR and tremor between the P15 and adult rats. These data indicate age-dependent differences in sensitivity to DLM. [ABSTRACT FROM AUTHOR]

Permethrin is a type I (noncyano) pyrethroid that induces tremors at high concentrations and increases acoustic startle responses (ASRs) in adult rodents, however its effects in young rats have been investigated to a limited extent. ASR and tremor were assessed in adult and postnatal day (P)15 Sprague-Dawley rats at oral doses of 60, 90, or 120 mg/kg over an 8 h period. Permethrin increased ASR in adults, regardless of dose, and 20% of the high-dose rats showed tremor at later time points. For the P15 rats all doses induced tremor at all time points, and ASR was increased at 2 h in the 90 and 120 mg/kg groups with a trend in the 60 mg/kg group compared with controls. The 60 mg/kg group showed increased ASR at 4 and 6 h, whereas the 90 mg/kg group showed no differences from the controls at these times. The 120 mg/kg group showed decreased ASR from 4- to 8-h posttreatment. P15 and adult rats both showed plasma and brain cis - and trans -permethrin increases after dosing. After the same dose of permethrin, P15 rats had greater cis - and trans -permethrin in brain and plasma compared with adults. P15 rats had an increased tremor response compared with adults even at comparable brain permethrin concentrations. For ASR, P15 rats responded sooner and showed a biphasic pattern ranging from increased to decreased response as a function of dose and time, unlike adults that only showed increases. Overall, young rats showed greater effects from permethrin compared with adults. [ABSTRACT FROM AUTHOR]

Methamphetamine (MA) alters dopamine markers and cognitive function in heavy users. In rodents, there are MA dosing regimens that induce concordant effects using repeated administration at spaced intervals. These regimens are effective but complicate experiments designed to disentangle the effects of the drug on different brain regions in relation to their cognitive effects because of treatment spacing. In an effort to simplify the model, we tested whether a single dose of MA could induce the same monoamine and cognitive effects as multiple, spaced dosing without affecting survival. Adult male Sprague-Dawley rats were treated with 40 mg/kg MA subcutaneously once and tested starting 2 weeks later. MA-treated rats showed deficits in egocentric navigation in Cincinnati water maze, in spatial navigation in the Morris water maze, and in choosing a consistent problem-solving strategy in the Star water maze when given the option to show a preference. MA-treated rats had persistent dopamine and serotonin reductions in the neostriatum and nucleus accumbens, and serotonin reductions in the hippocampus of the same magnitude as in repetitive treatment models. The data demonstrate that a single dose recapitulates the neurocognitive and monoamine effects of multiple-dose regimens, thereby simplifying the model of MA-induced neurotoxicity. [ABSTRACT FROM AUTHOR]

In utero methamphetamine (MA) exposure leads to a range of adverse effects, such as decreased attention, reduced working-memory capability, behavioral dysregulation, and spatial memory impairments in exposed children. In the current experiment, preweaning Sprague-Dawley rats-as a model of third trimester human exposure-were administered the spin trapping agent, N-tert-butyl-α-phenylnitrone (PBN), daily prior to MA. Rats were given 0 (SAL) or 40 mg/kg PBN prior to each MA dose (10 mg/kg, 4× per day) from postnatal day (P) 6-15. Littermates underwent Cincinnati water maze, Morris water maze, and radial water maze assessment beginning on P30 (males) or P60 (females). Males were also tested for conditioned contextual and cued freezing, while females were trained in passive avoidance. Findings show that, regardless of age/sex, neonatal MA induced deficits in all tests, except passive avoidance. PBN did not ameliorate these effects, but had a few minor effects. Taken together, MA induced learning deficits emerge early and persist, but the mechanism remains unknown. [ABSTRACT FROM AUTHOR]

Recent studies suggest that ultra-high dose rates of proton radiation (>40 Gy/s; FLASH) confer less toxicity to exposed healthy tissue and reduce cognitive decline compared with conventional radiation dose rates (~1 Gy/s), but further preclinical data are required to demonstrate this sparing effect. In this study, postnatal day 11 (P11) rats were treated with whole brain irradiation with protons at a total dose of 0, 5, or 8 Gy, comparing a conventional dose rate of 1 Gy/s vs. a FLASH dose rate of 100 Gy/s. Beginning on P64, rats were tested for locomotor activity, acoustic and tactile startle responses (ASR, TSR) with or without prepulses, novel object recognition (NOR; 4-object version), striatal dependent egocentric learning ([configuration A] Cincinnati water maze (CWM-A)), prefrontal dependent working memory (radial water maze (RWM)), hippocampal dependent spatial learning (Morris water maze (MWM)), amygdala dependent conditioned freezing, and the mirror image CWM [configuration B (CWM-B)]. All groups had deficits in the CWM-A procedure. Weight reductions, decreased center ambulation in the open-field, increased latency on day-1 of RWM, and deficits in CWM-B were observed in all irradiated groups, except the 5 Gy FLASH group. ASR and TSR were reduced in the 8 Gy FLASH group and day-2 latencies in the RWM were increased in the FLASH groups compared with controls. There were no effects on prepulse trials of ASR or TSR, NOR, MWM, or conditioned freezing. The results suggest striatal and prefrontal cortex are sensitive regions at P11 to proton irradiation, with reduced toxicity from FLASH at 5 Gy. [ABSTRACT FROM AUTHOR]

Latrophilin‐3 (LPHN3) is a brain specific G‐protein coupled receptor associated with increased risk of attention deficit hyperactivity disorder (ADHD) and cognitive deficits. CRISPR/Cas9 was used to generate a constitutive knockout (KO) rat of Lphn3 by deleting exon 3, based on human data that LPHN3 variants are associated with some cases of ADHD. Lphn3 KO rats are hyperactive with an attenuated response to ADHD medication and have cognitive deficits. Here, we tested KO, heterozygous (HET), and wildtype (WT) rats to determine if there was a gene‐dosage effect. We tested the rats in home‐cage activity starting at postnatal day (P)35 and P50, followed by tests of egocentric learning (Cincinnati water maze [CWM]), spatial learning (Morris water maze [MWM]), working memory (radial water maze [RWM]), incidental learning (novel object recognition [NOR]), acoustic startle response (ASR) habituation, tactile startle response (TSR) habituation, prepulse modification of acoustic startle, shuttle‐box passive avoidance, conditioned freezing, and a mirror image version of the CWM. KO and HET rats were hyperactive. KO and HET rats had egocentric (CWM) and spatial deficits (MWM), increased startle response, and KO rats showed less conditioned freezing on contextual and cued memory; there were no effects on working memory (RWM) or passive avoidance. The selective gene‐dosage effect in Lphn3 HET rats indicates that Lphn3 exhibits dominate expression on functions where it is most abundantly expressed (striatum, hippocampus) but not on behaviors mediated by regions of low expression. The data add further evidence to the impact of this synaptic protein on brain function and behavior. [ABSTRACT FROM AUTHOR]

Abstract: Methylphenidate (MPD) is the most prescribed drug for attention deficit hyperactivity disorder. Licit and illicit use also occurs during pregnancy, however the effects from this use on offspring development are unknown. To model late gestational exposure, Sprague–Dawley litters were treated with 0, 5, 10, 20, or 30mg/kg×4/day every 2h with MPD on postnatal days 11–20 (within-litter design; days chosen to be comparable to human third trimester brain development). During treatment, body weights were decreased in MPD-treated groups; weight recovery occurred in all but the MPD-30 group by start of testing. MPD-treated rats showed no changes in anxiety (elevated zero maze), swimming ability (straight channel swimming), or spatial learning/reference memory (Morris water maze). MPD does not appear to pose a risk to these CNS functions after exposure during a stage of rat development analogous to third trimester human brain development. [Copyright &y& Elsevier]

Abstract: The elevated plus and zero mazes (Plus and Zero, respectively) are used to assess behavior related to anxiety in rodents but direct comparisons of the two tests are lacking for rats. We compared the two methods in adult male Sprague–Dawley rats. Untreated rats in the Zero spent more time in open zones and exhibited more head dips than in the Plus whereas start latency and closed area entries were lower in the Zero than in the Plus. Diazepam (1mg/kg) exposure increased time in the open in both mazes. Restraint (60min prior to testing), yohimbine (2.5mg/kg), and caffeine (100mg/kg) had the opposite effect, significantly decreasing time spent in open zones in both mazes. No sexual dimorphism in behavior was seen in either maze in untreated rats. Although more open area time was evident in untreated animals in the Zero, after drug challenge both mazes detected anxiolytic and anxiogenic effects equally. Zero maze data can be analyzed directly because no center region exists; otherwise the two methods appear comparable following challenge. [ABSTRACT FROM AUTHOR]

Methamphetamine (MA) is widely abused and implicated in residual cognitive deficits. In rats, increases in plasma corticosterone and egocentric learning deficits are observed after a 1-day binge regimen of MA (10 mg/kg × 4 at 2-h intervals). The purpose of this experiment was to determine if adrenal inactivation during and following MA exposure would attenuate the egocentric learning deficits in the Cincinnati water maze (CWM). In the first experiment, the effects of adrenalectomy (ADX) or sham surgery (SHAM) on MA-induced neurotoxicity at 72 h were determined. SHAM-MA animals showed typical patterns of hyperthermia, whereas ADX-MA animals were normothermic. Both SHAM-MA- and ADX-MA-treated animals showed increased neostriatal glial fibrillary acidic protein and decreased monoamines in the neostriatum, hippocampus, and entorhinal cortex. In the second experiment, SHAM-MA- and ADX-MA-treated groups showed equivalently impaired CWM performance 2 weeks post-treatment (increased latencies, errors, and start returns) compared to SHAM-saline (SAL) and ADX-SAL groups with no effects on novel object recognition, elevated zero maze, or acoustic startle/prepulse inhibition. Post-testing, monoamine levels remained decreased in both MA-treated groups in all three brain regions, but were not as large as those observed at 72-h post-treatment. The data demonstrate that MA-induced learning deficits can be dissociated from drug-induced increases in plasma corticosterone or hyperthermia, but co-occur with dopamine and serotonin reductions. Synapse 64:773-785, 2010. © 2010 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Abstract: Mucopolysaccharidosis type I (MPS I) is one of the most common lysosomal storage diseases with progressive neurological dysfunction. To characterize the chronological behavioral profiles and identify the onset of functional deficits in a MPS I mouse model (IDUA−/−), we evaluated anxiety, locomotor behavior, startle, spatial learning and memory with mice at 2, 4, 6 and 8 months of age. In automated open-field test, IDUA−/− mice showed hypoactivity as early as 2 months of age and altered anxiety starting from 6 months of age during the initial exploratory phase, even though normal habituation was observed at all ages. In the marble-burying task, the anxiety-like compulsive behavior was normal in IDUA−/− mice at almost all tested ages, but significantly reduced in 8-month old male IDUA−/− mice which coincided with the rapid death of IDUA−/− males starting from 7 months of age. In the Morris water maze, IDUA−/− mice exhibited impaired proficient learning only at 4 months of age during the acquisition phase. Spatial memory deficits were observed in IDUA−/− mice during both 1 and 7 days probe trials at 4 and 8 months of age. The IDUA−/− mice performed normally in a novel object recognition task at younger ages until 8 months old when reduced visual cognitive memory retention was noted in the IDUA−/− mice. In addition, 8-month-old IDUA−/− mice failed to habituate to repeated open-field exposure, suggesting deficits in non-aversive and non-associative memory. In acoustic startle assessment, significantly more non-responders were found in IDUA−/− mice, but normal performance was seen in those that did show a response. These results presented a temporal evaluation of phenotypic behavioral dysfunctions in IDUA−/− mice from adolescence to maturity, indicating the impairments, with different ages of onset, in locomotor and anxiety-like compulsive behaviors, spatial learning and memory, visual recognition and short-term non-associative memory retention. This study would also provide guidelines for the experimental designs of behavioral evaluation on innovative therapies for the treatment of MPS type I. [Copyright &y& Elsevier]

Previous studies in rats showed that postnatal day (P)11–20 exposure to ±3,4-methylenedioxymethamphetamine (MDMA, ecstasy) causes learning and memory deficits in adulthood. The emergence and permanence of these learning deficits are currently unknown. This study was designed to investigate learning and memory deficits in adolescent (P30 or P40) and older (P180 or P360) rats exposed to MDMA from P11–20. Within each litter half the animals were exposed to MDMA (20 mg/kg) and half to saline (SAL) twice a day (8 h apart) from P11–20. In experiment (exp) 1, behavioral testing began on either P30 or P40, whereas in exp 2, testing began on either P180 or P360. Offspring were tested in the Cincinnati water maze (CWM), a test of path integration learning (2 trials/day for 5 days), and the Morris water maze (MWM) (three phases, with 5 days of 4 trials/day and a probe trial on the sixth day per phase). MDMA-treated rats took longer to find the platform and traveled a greater distance to find the platform at all ages tested in all phases of the MWM. MDMA-treated animals also spent less time in the target quadrant during probe trials. In the CWM, P30 and P40 animals took longer to find the goal and committed more errors in locating the goal, while P180 and P360 MDMA-treated animals performed similarly to SAL-treated animals. The data suggest that the spatial learning and memory deficits induced by MDMA are long lasting, while the path integration deficits recover over time. [ABSTRACT FROM AUTHOR]

Abstract: Background and purpose: Hypoxic–ischemic injury in term neonates remains a significant cause of long-term neurological morbidity. The post-natal day 10 (P10) mouse is accepted as a model for the term human. This study was designed to assess the relationships between the duration of hypoxia–ischemia (HI) on P10 and the structural and functional neurological deficits that appear in the adult mouse as a consequence. Methods: Post-natal day 10 129T2×C57Bl/6 F1 hybrid mice were subjected to 0, 45, 60 or 75 min of hypoxia–ischemia using the Rice–Vannucci model. Beginning on P50 these mice were tested over the next 8 weeks using zero maze, locomotor activity, novel object recognition, cued, hidden and reduced Morris water mazes, delayed probe trials and response to apomorphine injection. Brain weights and histology were obtained at the end of testing. Results: The degree of structural and behavioral abnormalities in adult mice correlated with the duration of hypoxia–ischemia on P10. Useful behavioral tests for separating adult mice according to duration of hypoxia–ischemia on P10 include locomotor activity, the Morris water mazes and response to apomorphine. We found cued “learning” persisted, although latencies increased, with increasing HI time while spatial learning decayed as a function of HI time. Severe HI injury involving the ventral hippocampus resulted in excessive locomotor activity. Conclusions: After correcting for motor deficits, there is evidence for persistence of “cued” learning but not spatial learning with increasing hypoxia–ischemia time on P10 in this model system. [Copyright &y& Elsevier]

Background: ±3,4-Methylenedioxymethamphetamine (MDMA) is a recreational drug that causes cognitive deficits in humans. A rat model for learning and memory deficits has not been established, although some cognitive deficits have been reported. Methods: Male Sprague-Dawley rats were treated with MDMA (15 mg/kg x 4 doses) or saline (SAL) (n = 20/treatment group) and tested in different learning paradigms: 1) path integration in the Cincinnati water maze (CWM), 2) spatial learning in the Morris water maze (MWM), and 3) novel object recognition (NOR). One week after drug administration, testing began in the CWM, then four phases of MWM, and finally NOR. Following behavioral testing, monoamine levels were assessed. Results: ±3,4-Methylenedioxymethamphetamine-treated rats committed more CWM errors than did SAL-treated rats. ±3,4-Methylenedioxymethamphetamine-treated animals were further from the former platform position during each 30-second MWM probe trial but showed no differences during learning trials with the platform present. There were no group differences in NOR. ±3,4-Methylenedioxymethamphetamine depleted serotonin in all brain regions and dopamine in the striatum. Conclusions: ±3,4-Methylenedioxymethamphetamine produced MWM reference memory deficits even after complex learning in the CWM, where deficits in path integration learning occurred. Assessment of path integration may provide a sensitive index of MDMA-induced learning deficits. [Copyright &y& Elsevier]

The effects of the dopamine D1 receptor antagonist R(+)-SCH-23390 and D2 receptor antagonist S(-)-eticlopride on d-methamphetamine-induced striatal monoamine reductions 72 h after treatment were investigated in relation to changes in body temperature. Rats were administered four 10-mg/kg doses of d-methamphetamine or saline with a 2-h interval between treatments; 0.5 mg/kg eticlopride or SCH-23390 was administered 15 min before each methamphetamine or saline injection. Two ambient temperature conditions were investigated: 24 and 33°C. Methamphetamine administered at 24°C induced hyperthermia and reduced striatal dopamine content by 73%; 0.5 mg/kg eticlopride or SCH-23390 administered in combination with methamphetamine at 24°C attenuated methamphetamine-induced hyperthermia and prevented significant reductions in dopamine content. At 33°C, eticlopride and SCH-23390 were ineffective in blocking methamphetamine-induced hyperthermia and dopamine content was reduced by 65% in the SCH-23390-methamphetamine group. By contrast, dopamine content was reduced by only 31% in the 33°C eticlopride-methamphetamine group. Thus, although the eticlopride-methamphetamine treatment combination at 33°C exhibited a hyperthermic response comparable to that seen with methamphetamine alone at 24°C, reductions in dopamine content were attenuated in the combination group compared with methamphetamine alone at 24°C. Serotonin changes showed similar attenuated reductions after SCH-23390 or eticlopride pretreatment at 24°C in combination with methamphetamine, but this attenuation was absent at 33°C. The dissociation of methamphetamine-induced striatal dopamine reduction and hyperthermia seen after eticlopride pretreatment suggests a dopamine D2 receptor mechanism in mediating methamphetamine-induced dopamine depletion. However this D2 mechanism does not apply to methamphetamine-induced striatal serotonin reductions. Synapse 56:84-93, 2005. © 2005 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

The investigation of methamphetamine exposure during neonatal development in rats has demonstrated that long-term spatial learning deficits are induced. A previous dose–response study showed that administration of 5 mg/kg methamphetamine, four times daily from postnatal days 11 to 20 produced these deficits, although the effects were not as severe as at higher doses of 10 or 15 mg/kg. This study examined concentrations of methamphetamine at or below 5 mg/kg given over the same period of time. Five different concentrations of methamphetamine (i.e., 5, 2.5, 1.25, 0.625, or 0) were administered every 2 h four times daily from postnatal days 11 to 20. Body weights, zero maze performance, and Morris water maze learning were examined. A dose-dependent decrease in body weight was observed during the period of methamphetamine administration and these lower weights continued throughout adulthood for the 5, 2.5, and 1.25 mg/kg concentrations, although the adult decreases were negligible. No differences were noted in the zero maze. In the Morris water maze during the acquisition period, dose-dependent differences in spatial orientation were seen, however non-dose related deficits were observed for other parameters. During the shifted platform phase (“reversal”), a similar dose-dependent difference in spatial orientation was observed, although no other effects were noted during this phase. Females performed worse than males regardless of treatment or the phase of learning in the Morris water maze. These data suggest that even lower doses of methamphetamine can alter learning and memory in adulthood, although with less consistent results than with doses higher than 5 mg/kg/dose. These data would caution against even casual use of methamphetamine by women during pregnancy since even low doses could alter the ability of the child to learn. [Copyright &y& Elsevier]

Previous studies have demonstrated that rats exposed to methamphetamine (MA) during the neonatal period display deficits in spatial learning and memory. The underlying correlates are unknown; therefore, this study was devised to determine whether neuronal changes occur in the dentate gyrus (DG), nucleus accumbens (NAcc) and cortex of adult rats exposed to 10 mg/kg MA administered four times daily from P11–20 using Golgi–Cox staining [Gibb, R. & Kolb, B. (1998) J. Neurosci. Meth., 79, 1–4]. The DG and NAcc demonstrated a decrease in the number of spines per neuron and the NAcc showed an associated decrease in dendritic length. Selective changes in cortex were observed because increased dendritic length in the parietal cortex occurred with no change in the number of spines, and no differences were noted for either dendritic length or spines in the medial frontal cortex. The data suggest a potential cause for the learning and memory deficits induced by neonatal MA exposure; however, the underlying mechanism that produces these neuronal changes is unknown. [ABSTRACT FROM AUTHOR]

Attention‐deficit/hyperactivity disorder (ADHD) a common neurodevelopmental disorder of childhood and often comorbid with other externalizing disorders (EDs). There is evidence that externalizing behaviors share a common genetic etiology. Recently, a genome‐wide, multigenerational sample linked variants in the Lphn3 gene to ADHD and other externalizing behaviors. Likewise, limited research in animal models has provided converging evidence that Lphn3 plays a role in EDs. This study examined the impact of Lphn3 deletion (i.e., Lphn3−/−) in rats on measures of behavioral control associated with externalizing behavior. Impulsivity was assessed for 30 days via a differential reinforcement of low rates (DRL) task and working memory evaluated for 25 days using a delayed spatial alternation (DSA) task. Data from both tasks were averaged into 5‐day testing blocks. We analyzed overall performance, as well as response patterns in just the first and last blocks to assess acquisition and steady‐state performance, respectively. "Positive control" measures on the same tasks were measured in an accepted animal model of ADHD–the spontaneously hypertensive rat (SHR). Compared with wildtype controls, Lphn3−/− rats exhibited deficits on both the DRL and DSA tasks, indicative of deficits in impulsive action and working memory, respectively. These deficits were less severe than those in the SHRs, who were profoundly impaired on both tasks compared with their control strain, Wistar‐Kyoto rats. The results provide evidence supporting a role for Lphn3 in modulating inhibitory control and working memory, and suggest additional research evaluating the role of Lphn3 in the manifestation of EDs more broadly is warranted. [ABSTRACT FROM AUTHOR]