Lack of Vesicular Zinc Does Not Affect the Behavioral Phenotype of Polyinosinic:Polycytidylic Acid-Induced Maternal Immune Activation Mice.

Zinc is important in neural and synaptic development and neuronal transmission. Within the brain, zinc transporter 3 (ZnT3) is essential for zinc uptake into vesicles. Loss of vesicular zinc has been shown to produce neurodevelopmental disorder (NDD)-like behavior, such as decreased social interaction and increased anxiety- and repetitive-like behavior. Maternal immune activation (MIA) has been identified as an environmental factor for NDDs, such as autism spectrum disorders (ASDs) and schizophrenia (SZ), in offspring, which occurs during pregnancy when the mother's immune system reacts to the exposure to viruses or infectious diseases. In this study, we investigated the interaction effect of a genetic factor [ZnT3 knockout (KO) mice] and an environmental factor (MIA). We induced MIA in pregnant female (dams) mice during mid-gestation, using polyinosinic:polycytidylic acid (polyI:C), which mimics a viral infection. Male and female ZnT3 KO and wild-type (WT) offspring were tested in five behavioral paradigms: Ultrasonic Vocalizations (USVs) at postnatal day 9 (P9), Open Field Test, Marble Burying Test, three-Chamber Social Test, and Pre-pulse Inhibition (PPI) in adulthood (P60-75). Our results indicate that loss of vesicular zinc does not result in enhanced ASD- and SZ-like phenotype compared to WT, nor does it show a more pronounced phenotype in male ZnT3 KO compared to female ZnT3 KO. Finally, MIA offspring demonstrated an ASD- and SZ-like phenotype only in specific behavioral tests: increased calls emitted in USVs and fewer marbles buried. Our results suggest that there is no interaction between the loss of vesicular zinc and MIA induction in the susceptibility to developing an ASD- and SZ-like phenotype.

Examination of Zinc in the Circadian System.

Zinc is a trace element that is essential for a large number of biological and biochemical processes in the body. In the nervous system zinc is packaged into synaptic vesicles by the ZnT3 transporter, and synaptic release of zinc can influence the activity of postsynaptic cells, either directly through its own cognate receptors, or indirectly by modulating activation of receptors for other neurotransmitters. Here, we explore the anatomical and functional aspects of zinc in the circadian system. Melanopsin-containing retinal ganglion cells in the mouse retina were found to colocalize ZnT3, indicating that they can release zinc at their synaptic targets. While the master circadian clock in the hamster suprachiasmatic nucleus (SCN) was found to contain, at best, sparse zincergic input, the intergeniculate leaflet (IGL) of hamsters and mice were found to have prominent zincergic input. Levels of zinc in these areas were not affected by time of day. Additionally, IGL zinc staining persisted following enucleation, indicating other prominent sources of zinc instead of, or in addition to, the retina. Neither enhancement nor chelation of free zinc at either the SCN or IGL altered circadian responses to phase-shifting light in hamsters. Finally, entrainment, free-running, and circadian responses to light were explored in mice lacking the ZnT3 gene. In every aspect explored, the ZnT3 knockout mice were not significantly different from their wildtype counterparts. These findings highlight the presence of zinc in areas critical for circadian functioning but have yet to identify a role for zinc in these areas.

Effects of enriched housing on the neuronal morphology of mice that lack zinc transporter 3 (ZnT3) and vesicular zinc.

In the central nervous system, certain neurons store zinc within the synaptic vesicles of their axon terminals. This vesicular zinc can then be released in an activity-dependent fashion as an intercellular signal. The functions of vesicular zinc are not entirely understood, but evidence suggests that it is important for some forms of experience-dependent plasticity in the brain. The ability of neurons to store and release vesicular zinc is dependent on expression of the vesicular zinc transporter, ZnT3. Here, we examined the neuronal morphology of mice that lack ZnT3. Brains were collected from mice housed under standard laboratory conditions and from mice housed in enriched environments - large, multilevel enclosures with running wheels, numerous objects and tunnels, and a greater number of cage mates. Golgi-Cox staining was used to visualize neurons for analysis of dendritic length and dendritic spine density. Neurons were analyzed from the barrel cortex, striatum, basolateral amygdala, and hippocampus (CA1). ZnT3 knockout mice, relative to wild type mice, exhibited increased basal dendritic length in the layer 2/3 pyramidal neurons of barrel cortex, independently of housing condition. Environmental enrichment decreased apical dendritic length in these same neurons and increased dendritic spine density on striatal medium spiny neurons. Elimination of ZnT3 did not modulate any of the effects of enrichment. Our results provide no evidence that vesicular zinc is required for the experience-dependent changes that occur in response to environmental enrichment. They are consistent, however, with recent reports suggesting increased cortical volume in ZnT3 knockout mice.

Behavioral characterization of female zinc transporter 3 (ZnT3) knockout mice.

Zinc is an important element in all cells of the body, having structural, enzymatic, and regulatory functions. In some neurons, zinc is loaded into synaptic vesicles by zinc transporter 3 (ZnT3) and released into the synaptic cleft, where it can modulate neuronal function. ZnT3 knockout (KO) mice lack ZnT3 and thus lack synaptic zinc. Previous studies have examined the behavioral phenotype of ZnT3 KO mice, mostly using mixed-sex or male-only groups. In the present study we focused specifically on the behavior of female ZnT3 KO mice (2-3 months old). An extensive battery of tests was administered to assess sensorimotor and cognitive behaviours, as well as to examine for a possible schizophrenia-like phenotype. ZnT3 KO mice performed similarly to wild type controls in the majority of tests. However, they were less accurate in the skilled reach task, suggesting impaired skilled motor learning, and faster to descend a vertical pole. ZnT3 KO mice were also slower in the open field and made fewer chamber entries in the social preference test, suggesting decreased exploratory locomotion. No differences were observed in the Morris water task or fear conditioning test. This is the first study to show a behavioural phenotype specifically for female ZnT3 KO mice. Comparing our results to previous studies, it appears that there may be sex-specific effects of eliminating ZnT3. Female ZnT3 KO mice exhibit abnormalities in locomotion and at skilled motor learning, but we were unable to detect spatial or fear learning deficits previously described in male ZnT3 KO mice.