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Botterill, J. J.

Publications and source records attributed to Botterill, J. J..

9 recordsLinked to original sources
bioRxiv2024

Field EPSPs of dentate gyrus granule cells studied by selective optogenetic activation of hilar mossy cells in hippocampal slices.

Glutamatergic dentate gyrus (DG) mossy cells (MCs) innervate the primary DG cell type, granule cells (GCs). Numerous MC synapses are on GC proximal dendrites in the inner molecular layer (IML). However, field recordings of the GC excitatory postsynaptic potential (fEPSPs) have not been used to study this pathway selectively. Here we describe methods to selectively activate MC axons in the IML using mice with Cre recombinase expressed in MCs. Slices were made after injecting adeno-associated virus (AAV) encoding channelrhodopsin (ChR2) in the DG. In these slices, we show that fEPSPs could be recorded reliably in the IML in response to optogenetic stimulation of MC axons. Furthermore, fEPSPs were widespread across the septotemporal axis. However, fEPSPs were relatively weak because they were small in amplitude and did not elicit a significant population spike in GCs. They also showed little paired pulse facilitation. We confirmed the extracellular findings with patch clamp recordings of GCs despite different recording chambers and other differences in methods. Together the results provide a simple method for studying MC activation of GCs and add to the evidence that this input is normally weak but widespread across the GC population. KEY POINTSO_LIWe describe a method to activate the MC input to GCs selectively using optogenetics in hippocampal slices C_LIO_LIMC excitation is weakly excitatory but so common among GCs that a field EPSP is generated at the site of MC synapses on GCs C_LIO_LIMC excitation of GCs is consistent across the septotemporal axis and contralaterally C_LIO_LIUsing the characteristics of optogenetically-evoked fEPSPs, electrical stimulation of the MC input to GCs can be optimized. C_LI

neuroscience↗
bioRxiv2024

Increased excitability of dentate gyrus mossy cells occurs early in life in the Tg2576 model of Alzheimer's disease.

INTRODUCTIONHyperexcitability in Alzheimers disease (AD) emerge early and contribute to disease progression. The dentate gyrus (DG) is implicated in hyperexcitability in AD. We hypothesized that mossy cells (MCs), regulators of DG excitability, contribute to early hyperexcitability in AD. Indeed, MCs generate hyperexcitability in epilepsy. METHODSUsing the Tg2576 model and WT mice ([~]1month-old), we compared MCs electrophysiologically, assessed c-Fos activity marker, A{beta} expression and mice performance in a hippocampal-dependent memory task. RESULTSTg2576 MCs exhibit increased spontaneous excitatory events and decreased inhibitory currents, increasing the charge transfer excitation/inhibition ratio. Tg2576 MC intrinsic excitability was enhanced, and showed higher c-Fos, intracellular A{beta} expression, and axon sprouting. Granule cells only showed changes in synaptic properties, without intrinsic changes. The effects occurred before a memory task is affected. DISCUSSIONEarly electrophysiological and morphological alterations in Tg2576 MCs are consistent with enhanced excitability, suggesting an early role in DG hyperexcitability and AD pathophysiology. HIGHLIGHTS{circ} MCs from 1 month-old Tg2576 mice had increased spontaneous excitatory synaptic input. {circ}Tg2576 MCs had reduced spontaneous inhibitory synaptic input. {circ}Several intrinsic properties were abnormal in Tg2576 MCs. {circ}Tg2576 GCs had enhanced synaptic excitation but no changes in intrinsic properties. {circ}Tg2576 MCs exhibited high c-Fos expression, soluble A{beta} and axonal sprouting.

neuroscience↗
bioRxiv2023

Increasing adult neurogenesis protects mice from epilepsy.

ABSTRACTNeurogenesis occurs in the adult brain in the hippocampal dentate gyrus, an area that contains neurons which are vulnerable to insults and injury, such as severe seizures. Previous studies showed that increasing adult neurogenesis reduced neuronal damage after these seizures. Because the damage typically is followed by chronic life-long seizures (epilepsy), we asked if increasing adult-born neurons would prevent epilepsy. Adult-born neurons were selectively increased by deleting the pro-apoptotic gene Bax from Nestin-expressing progenitors. Tamoxifen was administered at 6 weeks of age to conditionally delete Bax in Nestin-CreERT2Baxfl/fl mice. Six weeks after tamoxifen administration, severe seizures (status epilepticus; SE) were induced by injection of the convulsant pilocarpine. After mice developed epilepsy, seizure frequency was quantified for 3 weeks. Mice with increased adult-born neurons exhibited fewer chronic seizures. Postictal depression was reduced also. These results were primarily in female mice, possibly because they were the more affected by Bax deletion than males, consistent with sex differences in Bax. The female mice with enhanced adult-born neurons also showed less neuronal loss of hilar mossy cells and hilar somatostatin-expressing neurons than wild type females or males, which is notable because these two hilar cell types are implicated in epileptogenesis. The results suggest that selective Bax deletion to increase adult-born neurons can reduce experimental epilepsy, and the effect shows a striking sex difference. The results are surprising in light of past studies showing that suppressing adult-born neurons can also reduce chronic seizures.

neuroscience↗
bioRxiv2023

Choline supplementation in early life improves and low levels of choline can impair outcomes in a mouse model of Alzheimer's disease.

Maternal choline supplementation (MCS) improves cognition in Alzheimers disease (AD) models. However, effects of MCS on neuronal hyperexcitability in AD are unknown. We investigated effects of MCS in a well-established mouse model of AD with hyperexcitability, the Tg2576 mouse. The most common type of hyperexcitability in Tg2576 mice are generalized EEG spikes (interictal spikes; IIS). IIS also are common in other mouse models and occur in AD patients. Im mouse models, hyperexcitability is also reflected by elevated expression of the transcription factor {Delta}FosB in the granule cells (GCs) of the dentate gyrus (DG), which are the principal cell type. Therefore we studied {Delta}FosB expression in GCs. We also studied the the neuronal marker NeuN within hilar neurons of the DG because other studies have reduced NeuN protein expression is a sign of oxidative stress or other pathology. This is potentially important because hilar neurons regulate GC excitability. Tg2576 breeding pairs received a diet with a relatively low, intermediate or high concentration of choline. After weaning, all mice received the intermediate diet. In offspring of mice fed the high choline diet, IIS frequency declined, GC {Delta}FosB expression was reduced, and NeuN expression was restored. Using the novel object location task, spatial memory improved. In contrast, offspring exposed to the relatively low choline diet had several adverse effects, such as increased mortality. They had the weakest hilar NeuN immunoreactivity and greatest GC {Delta}FosB protein expression. However, their IIS frequency was low, which was surprising. The results provide new evidence that a diet high in choline in early life can improve outcomes in a mouse model of AD, and relatively low choline can have mixed effects. This is the first study showing that dietary choline can regulate hyperexcitability, hilar neurons, {Delta}FosB and spatial memory in an animal model of AD.

neuroscience↗
bioRxiv2023

Dorsal peduncular cortex activity modulates affective behaviors in mice

The medial prefrontal cortex (mPFC) is critical to cognitive and emotional function and underlies many neuropsychiatric disorders, including mood, fear and anxiety disorders. In rodents, disruption of mPFC activity affects anxiety- and depression-like behavior, with specialized contributions from its subdivisions. The rodent mPFC is divided into the dorsomedial prefrontal cortex (dmPFC), spanning the anterior cingulate cortex (ACC) and dorsal prelimbic cortex (PL), and the ventromedial prefrontal cortex (vmPFC), which includes the ventral PL, infralimbic cortex (IL), and in some studies the dorsal peduncular cortex (DP) and dorsal tenia tecta (DTT). The DP/DTT have recently been implicated in the regulation of stress- induced sympathetic responses via projections to the hypothalamus. While many studies implicate the PL and IL in anxiety-, depression-like and fear behavior, the contribution of the DP/DTT to affective and emotional behavior remains unknown. Here, we used chemogenetics and optogenetics to bidirectionally modulate DP/DTT activity and examine its effects on affective behaviors, fear and stress responses in C57BL/6J mice. Acute chemogenetic activation of DP/DTT significantly increased anxiety-like behavior in the open field and elevated plus maze tests, as well as passive coping in the tail suspension test. DP/DTT activation also led to an increase in serum corticosterone levels and facilitated auditory fear extinction learning and retrieval. Activation of DP/DTT projections to the dorsomedial hypothalamus (DMH) acutely decreased freezing at baseline and during extinction learning, but did not alter affective behavior. These findings point to the DP/DTT as a new regulator of affective behavior and fear extinction in mice.

neuroscience↗
bioRxiv2021

Off-target expression of Cre-dependent adeno-associated viruses in wild type C57BL/6J mice

Adeno-associated viruses (AAVs) are a commonly used tool in neuroscience to efficiently label, trace, and/or manipulate neuronal populations. Highly specific targeting can be achieved through recombinase-dependent AAVs in combination with transgenic rodent lines that express Cre-recombinase in specific cell types. Visualization of viral expression is typically achieved through fluorescent reporter proteins (e.g., GFP or mCherry) packaged within the AAV genome. Although non-amplified fluorescence is usually sufficient to observe viral expression, immunohistochemical amplification of the fluorescent reporter is routinely used to improve viral visualization. In the present study, Cre-dependent AAVs were injected into the hippocampus and cortex of wild-type C57BL/6J mice. While we observed weak but consistent non-amplified off-target DIO expression in C57BL/6J mice, antibody amplification of the GFP or mCherry reporter revealed extensive Cre-independent viral expression. Off-target expression of DIO constructs in wild-type C57BL/6J mice occurred independent of vendor, AAV serotype or promoter. We also evaluated whether Cre-independent expression had functional effects via Designer Receptors Exclusively Activated by Designer Drugs (DREADDs). The DREADD agonist C21 had no effect on contextual fear conditioning or cFos expression in DIO-hM3Dq-mCherry+ cells of C57BL/6J mice. Taken together, our results indicate that DIO constructs have considerable off-target expression in wild type subjects. Our findings are particularly important for the design of experiments featuring sensitive systems and/or quantitative measurements that could be negatively impacted by off-target expression. Significance StatementAdeno-associated viruses (AAV) are widely used in neuroscience because of their safety and ease of use. Combined with specific promoters, Cre/loxP, and stereotaxic injections, highly specific targeting of cells and circuits within the brain can be achieved. In the present study we injected Cre-dependent AAVs into wild-type C57BL/6J mice and found considerable Cre-independent viral expression of AAVs encoding mCherry, GFP, or hM3Dq following immunohistochemical amplification of the fluorescent reporter protein. Importantly, we observed no functional effects of the Cre-independent expression in the hippocampus, as C21 had no detectable effect on DIO-hM3Dq-mCherry infected neurons in C57BL/6J mice. Given the widespread use of DIO rAAVs by the neuroscience community, our data supports careful consideration when using DIO constructs in control animals.

neuroscience↗
bioRxiv2020

Dorsal and ventral mossy cells differ in their long-range axonal projections throughout the dentate gyrus of the mouse hippocampus

Glutamatergic hilar mossy cells (MCs) have axons that terminate both near and far from their cell body but stay within the DG, making synapses in the inner molecular layer primarily. The long-range axons are considered the primary projection, and extend throughout the DG ipsilateral to the soma, and project to the contralateral DG. The specificity of long-range MC axons for the inner molecular layer (IML) has been considered to be a key characteristic of the DG. In the present study we made the surprising finding that dorsal MC axons are an exception to this rule. We used two mouse lines that allow for Cre-dependent viral labeling of MCs and their axons: dopamine receptor d2 (Drd2-Cre) and calcitonin receptor-like receptor (Crlr-Cre). A single viral injection into the dorsal DG to label dorsal MCs resulted in labeling of MC axons in both the IML and middle molecular layer (MML). Interestingly, this broad termination of MC axons applied to all long-range axons. In contrast, long-range axons of ventral MCs mainly terminated in the IML, consistent with the literature. Taken together, these results suggest that dorsal and ventral MCs differ significantly in their axonal projections, and the difference is primarily in their long-range projections. Since those projections are thought to terminate primarily on GCs, the results suggest a dorsal-ventral difference in MC activation of GCs. The surprising difference in dorsal and ventral MC projections should therefore be considered when evaluating dorsal-ventral differences in DG function.

neuroscience↗
bioRxiv2020

Bidirectional regulation of cognitive and anxiety-like behaviors by dentate gyrus mossy cells in male and female mice

The dentate gyrus (DG) of the hippocampus is important for cognitive and affective behaviors. However, the circuits underlying these behaviors are unclear. DG mossy cells (MCs) have been a focus of attention because of their excitatory synapses on the primary DG cell type, granule cells (GCs). However, MCs also activate DG GABAergic neurons which inhibit GCs. We took advantage of specific methods and a gain- and loss-of function strategy with Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) to study MCs in diverse behaviors. Using this approach, manipulations of MCs could bidirectionally regulate behavior. The results suggest that inhibiting MCs can reduce anxiety-like behavior and improve cognitive performance. However, not all cognitive or anxiety-related behaviors were influenced, suggesting specific roles of MCs in some but not all types of cognition and anxiety. Notably, several behaviors showed sex-specific effects, with females often showing more pronounced effects than the males. We also used the immediate early gene c-Fos to address whether DREADDs bidirectionally regulated MC or GC activity. We confirmed excitatory DREADDs increased MC c-Fos. However, there was no change in GC c-Fos, consistent with MC activation leading to GABAergic inhibition of GCs. In contrast, inhibitory DREADDs led to a large increase in GC c-Fos, consistent with a reduction in MC excitation of GABAergic neurons, and reduced inhibition of GCs. Taken together, these results suggest that MCs regulate anxiety and cognition in specific ways. We also raise the possibility that cognitive performance may be improved by reducing anxiety. SIGNIFICANCE STATEMENTThe dentate gyrus (DG) has many important cognitive roles as well as being associated with affective behavior. This study addressed how a glutamatergic DG cell type called mossy cells (MCs) contributes to diverse behaviors, which is timely because it is known that MCs regulate the activity of the primary DG cell type, granule cells (GCs), but how MC activity influences behavior is unclear. We show, surprisingly, that activating MCs can lead to adverse behavioral outcomes, and inhibiting MCs have an opposite effect. Importantly, the results appeared to be task-dependent and showed that testing both sexes was important. Additional experiments indicated what MC and GC circuitry was involved. Taken together, the results suggest how MCs influence behaviors that involve the DG.

neuroscience↗