The laterodorsal tegmentum and seizure regulation: Revisiting the evidence.

Electrical deep brain stimulation (DBS) is now a routine treatment option for patients suffering from medically refractory epilepsy. DBS of the anterior nucleus of the thalamus (ANT) has proven to be effective but, despite its success, few patients experience complete cessation of seizure activity. However, improving the therapy is challenging because the mechanism underlying its action remains largely unknown. One angle on improving the effectiveness of ANT stimulation is to better understand the various anatomic regions that send projections to and through this area. Here, the authors utilized a connectomic atlas of the mouse brain to better understand the regions projecting to the ANT and were particularly interested by the presence of robust cholinergic projections from the laterodorsal tegmentum (LDT). A subsequent review of the literature resulted in limited studies, which presented convincing evidence supporting this region's role in seizure control present in acute rodent models of epilepsy. It is thus the purpose of this paper to encourage further research into the role of the LDT on seizure mitigation, with mechanistic effects likely stemming from its cholinergic projections to the ANT. While previous studies have laid a firm foundation supporting the role of this region in modulation of seizure activity, modern scientific methodology has yet to be applied to further elucidate the mechanisms and potential benefits associated with LDT stimulation in the epileptic population.

The mesopontine tegmentum in reward and aversion: From cellular heterogeneity to behaviour.

The mesopontine tegmentum, comprising the pedunculopontine tegmentum (PPN) and the laterodorsal tegmentum (LDT), is intricately connected to various regions of the basal ganglia, motor systems, and limbic systems. The PPN and LDT can regulate the activity of different brain regions of these target systems, and in this way are in a privileged position to modulate motivated behaviours. Despite recent findings, the PPN and LDT have been largely overlooked in discussions about the neural circuits associated with reward and aversion. This review aims to provide a timely and comprehensive resource on past and current research, highlighting the PPN and LDT's connectivity and influence on basal ganglia and limbic, and motor systems. Seminal studies, including lesion, pharmacological, and optogenetic/chemogenetic approaches, demonstrate their critical roles in modulating reward/aversive behaviours. The review emphasizes the need for further investigation into the associated cellular mechanisms, in order to clarify their role in behaviour and contribution for different neuropsychiatric disorders.

Sex-dependent neuronal effects of α-synuclein reveal that GABAergic transmission is neuroprotective of sleep-controlling neurons.

BACKGROUND: Sleep disorders (SDs) are a symptom of the prodromal phase of neurodegenerative disorders that are mechanistically linked to the protein α-synuclein (α-syn) including Parkinson's disease (PD). SDs during the prodromal phase could result from neurodegeneration induced in state-controlling neurons by accumulation of α-syn predominant early in the disease, and consistent with this, we reported the monomeric form of α-syn (monomeric α-syn; α-synM) caused cell death in the laterodorsal tegmental nucleus (LDT), which controls arousal as well as the sleep and wakefulness state. However, we only examined the male LDT, and since sex is considered a risk factor for the development of α-syn-related diseases including prodromal SDs, the possibility exists of sex-based differences in α-synM effects. Accordingly, we examined the hypothesis that α-synM exerts differential effects on membrane excitability, intracellular calcium, and cell viability in the LDT of females compared to males. METHODS: Patch clamp electrophysiology, bulk load calcium imaging, and cell death histochemistry were used in LDT brain slices to monitor responses to α-synM and effects of GABA receptor acting agents. RESULTS: Consistent with our hypothesis, we found differing effects of α-synM on female LDT neurons when compared to male. In females, α-synM induced a decrease in membrane excitability and heightened reductions in intracellular calcium, which were reliant on functional inhibitory acid transmission, as well as decreased the amplitude and frequency of spontaneous excitatory postsynaptic currents (sEPSCs) with a concurrent reduction in action potential firing rate. Cell viability studies showed higher α-synM-mediated neurodegeneration in males compared to females that depended on inhibitory amino acid transmission. Further, presence of GABA receptor agonists was associated with reduced cell death in males. CONCLUSIONS: When taken together, we conclude that α-synM induces a sex-dependent effect on LDT neurons involving a GABA receptor-mediated mechanism that is neuroprotective. Understanding the potential sex differences in neurodegenerative processes, especially those occurring early in the disease, could enable implementation of sex-based strategies to identify prodromal PD cases, and promote efforts to illuminate new directions for tailored treatment and management of PD.

An inhibitory brainstem input to dopamine neurons encodes nicotine aversion.

Nicotine stimulates the dopamine (DA) system, which is essential for its rewarding effect. Nicotine is also aversive at high doses; yet, our knowledge about nicotine's dose-dependent effects on DA circuits remains limited. Here, we demonstrate that high doses of nicotine, which induce aversion-related behavior in mice, cause biphasic inhibitory and excitatory responses in VTA DA neurons that can be dissociated by distinct projections to lateral and medial nucleus accumben subregions, respectively. Guided by computational modeling, we performed a pharmacological investigation to establish that inhibitory effects of aversive nicotine involve desensitization of α4ß2 and activation of α7 nicotinic acetylcholine receptors. We identify α7-dependent activation of upstream GABA neurons in the laterodorsal tegmentum (LDT) as a key regulator of heterogeneous DA release following aversive nicotine. Finally, inhibition of LDT GABA terminals in VTA prevents nicotine aversion. Together, our findings provide a mechanistic circuit-level understanding of nicotine's dose-dependent effects on reward and aversion.

Prenatal nicotine alters development of the laterodorsal tegmentum: Possible role for attention-deficit/hyperactivity disorder and drug dependence.

As we cycle between the states of wakefulness and sleep, a bilateral cholinergic nucleus in the pontine brain stem, the laterodorsal tegmentum (LDT), plays a critical role in controlling salience processing, attention, behavioral arousal, and electrophysiological signatures of the sub- and microstates of sleep. Disorders involving abnormal alterations in behavioral and motivated states, such as drug dependence, likely involve dysfunctions in LDT signaling. In addition, as the LDT exhibits connectivity with the thalamus and mesocortical circuits, as well as receives direct, excitatory input from the prefrontal cortex, a role for the LDT in cognitive symptoms characterizing attention-deficit/hyperactivity disorder (ADHD) including impulsivity, inflexibility, and dysfunctions of attention is suggested. Prenatal nicotine exposure (PNE) is associated with a higher risk for later life development of drug dependence and ADHD, suggesting alteration in development of brain regions involved in these behaviors. PNE has been shown to alter glutamate and cholinergic signaling within the LDT. As glutamate and acetylcholine are major excitatory mediators, these alterations would likely alter excitatory output to target regions in limbic motivational circuits and to thalamic and cortical networks mediating executive control. Further, PNE alters neuronal development and transmission within prefrontal cortex and limbic areas that send input to the LDT, which would compound effects of differential processing within the PNE LDT. When taken together, alterations in signaling in the LDT are likely to play a role in negative behavioral outcomes seen in PNE individuals, including a heightened risk of drug dependence and ADHD behaviors.

The organization of cholinergic projections in the visual thalamus of the mouse.

Cholinergic projections from the brainstem serve as important modulators of activity in visual thalamic nuclei such as the dorsal lateral geniculate nucleus (dLGN). While these projections have been studied in several mammals, a comprehensive examination of their organization in the mouse is lacking. We used the retrograde transport of viruses or cholera toxin subunit B (CTB) injected in the dLGN, immunocytochemical labeling with antibodies against choline acetyltransferase (ChAT), brain nitric oxide synthase (BNOS), and vesicular acetylcholine transporter (VAChT), ChAT-Cre mice crossed with a reporter line (Ai9), as well as brainstem virus injections in ChAT-Cre mice to examine the pattern of thalamic innervation from cholinergic neurons in the pedunculopontine tegmental nucleus (PPTg), laterodorsal tegmental nucleus (LDTg), and the parabigeminal nucleus (PBG). Retrograde tracing demonstrated that the dLGN receives input from the PPTg, LDTg, and PBG. Viral tracing in ChAT-Cre mice and retrograde tracing combined with immunocytochemistry revealed that many of these inputs originate from cholinergic neurons in the PBG and PPTg. Most notable was an extensive cholinergic projection from the PBG which innervated most of the contralateral dLGN, with an especially dense concentration in the dorsolateral shell, as well as a small region in the dorsomedial pole of the ipsilateral dLGN. The PPTg was found to provide a sparse somewhat diffuse innervation of the ipsilateral dLGN. Neurons in the PPTg co-expressed ChAT, BNOS, and VAChT, whereas PBG neurons expressed ChAT, but not BNOS or VAChT. These results highlight the presence of distinct cholinergic populations that innervate the mouse dLGN.

Brain-wide Mapping of Mono-synaptic Afferents to Different Cell Types in the Laterodorsal Tegmentum.

The laterodorsal tegmentum (LDT) is a brain structure involved in distinct behaviors including arousal, reward, and innate fear. How environmental stimuli and top-down control from high-order sensory and limbic cortical areas converge and coordinate in this region to modulate diverse behavioral outputs remains unclear. Using a modified rabies virus, we applied monosynaptic retrograde tracing to the whole brain to examine the LDT cell type specific upstream nuclei. The LDT received very strong midbrain and hindbrain afferents and moderate cortical and hypothalamic innervation but weak connections to the thalamus. The main projection neurons from cortical areas were restricted to the limbic lobe, including the ventral orbital cortex (VO), prelimbic, and cingulate cortices. Although different cell populations received qualitatively similar inputs, primarily via afferents from the periaqueductal gray area, superior colliculus, and the LDT itself, parvalbumin-positive (PV+) GABAergic cells received preferential projections from local LDT neurons. With regard to the different subtypes of GABAergic cells, a considerable number of nuclei, including those of the ventral tegmental area, central amygdaloid nucleus, and VO, made significantly greater inputs to somatostatin-positive cells than to PV+ cells. Diverse inputs to the LDT on a system-wide level were revealed.

Impairments in laterodorsal tegmentum to VTA projections underlie glucocorticoid-triggered reward deficits.

Ventral tegmental area (VTA) activity is critical for reward/reinforcement and is tightly modulated by the laterodorsal tegmentum (LDT). In utero exposure to glucocorticoids (iuGC) triggers prominent motivation deficits but nothing is known about the impact of this exposure in the LDT-VTA circuit. We show that iuGC-rats have long-lasting changes in cholinergic markers in the LDT, together with a decrease in LDT basal neuronal activity. Interestingly, upon LDT stimulation, iuGC animals present a decrease in the magnitude of excitation and an increase in VTA inhibition, as a result of a shift in the type of cells that respond to the stimulus. In agreement with LDT-VTA dysfunction, we show that iuGC animals present motivational deficits that are rescued by selective optogenetic activation of this pathway. Importantly, we also show that LDT-VTA optogenetic stimulation is reinforcing, and that iuGC animals are more susceptible to the reinforcing properties of LDT-VTA stimulation.

Intra-ventral tegmental area microinjections of urotensin II modulate the effects of cocaine.

Although the peptide urotensin II (UII) has well studied direct actions on the cardiovascular system, the UII receptor (UIIR) is expressed by neurons of the hindbrain. Specifically, the UIIR is expressed by the cholinergic neurons of the laterodorsal tegmentum (LDTg) and the pedunculopontine tegmentum (PPTg). These neurons send axons to the ventral tegmental area (VTA), for which the PPTg and LDTg are the sole source of acetylcholine. Therefore, it was hypothesized that UIIR activation within the VTA would modulate reward-related behaviors, such as cocaine-induced drug seeking. Intra-VTA microinjections of UII at high concentrations (1 nmole) established conditioned place preference (CPP), but also blocked cocaine-mediated CPP (10 mg/kg). When rats received systemic sub-effectual doses of cocaine (7.5 mg/kg) with intra-VTA injections of 1 or 10 pmole of UII CPP was formed. Furthermore, the second endogenous ligand for the UIIR, urotensin II-related peptide, had the same effect at the 10 pmole dose. The effects of low doses of UII were blocked by pretreatment with the UIIR antagonist SB657510. Furthermore, it was found that intra-VTA UII (10 pmole) further increased cocaine-mediated (7.5 mg/kg) rises in electrically evoked dopamine in the nucleus accumbens. Our study has found that activation of VTA-resident UIIR produces observable behavioral changes in rats, and that UIIR is able to modulate the effects of cocaine. In addition, it was found that UIIR activation within the VTA can potentiate cocaine-mediated neurochemical effects. Therefore, the coincident activation of the UII-system and cocaine administration may increase the liability for drug taking behavior.

The cholinergic agonist carbachol increases the frequency of spontaneous GABAergic synaptic currents in dorsal raphe serotonergic neurons in the mouse.

Dorsal raphe nucleus (DRN) serotonin (5-HT) neurons play an important role in feeding, mood control and stress responses. One important feature of their activity across the sleep-wake cycle is their reduced firing during rapid-eye-movement (REM) sleep which stands in stark contrast to the wake/REM-on discharge pattern of brainstem cholinergic neurons. A prominent model of REM sleep control posits a reciprocal interaction between these cell groups. 5-HT inhibits cholinergic neurons, and activation of nicotinic receptors can excite DRN 5-HT neurons but the cholinergic effect on inhibitory inputs is incompletely understood. Here, in vitro, in DRN brain slices prepared from GAD67-GFP knock-in mice, a brief (3 min) bath application of carbachol (50 µM) increased the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) in GFP-negative, putative 5-HT neurons but did not affect miniature (tetrodotoxin-insensitive) IPSCs. Carbachol had no direct postsynaptic effect. Thus, carbachol likely increases the activity of local GABAergic neurons which synapse on 5-HT neurons. Removal of dorsal regions of the slice including the ventrolateral periaqueductal gray (vlPAG) region where GABAergic neurons projecting to the DRN have been identified, abolished the effect of carbachol on sIPSCs whereas the removal of ventral regions containing the oral region of the pontine reticular nucleus (PnO) did not. In addition, carbachol directly excited GFP-positive, GABAergic vlPAG neurons. Antagonism of both muscarinic and nicotinic receptors completely abolished the effects of carbachol. We suggest cholinergic neurons inhibit DRN 5-HT neurons when acetylcholine levels are lower i.e. during quiet wakefulness and the beginning of REM sleep periods, in part via excitation of muscarinic and nicotinic receptors located on local vlPAG and DRN GABAergic neurons. Higher firing rates or burst firing of cholinergic neurons associated with attentive wakefulness or phasic REM sleep periods leads to excitation of 5-HT neurons via the activation of nicotinic receptors located postsynaptically and presynaptically on excitatory afferents.

Reward and aversion in a heterogeneous midbrain dopamine system.

The ventral tegmental area (VTA) is a heterogeneous brain structure that serves a central role in motivation and reward processing. Abnormalities in the function of VTA dopamine (DA) neurons and the targets they influence are implicated in several prominent neuropsychiatric disorders including addiction and depression. Recent studies suggest that the midbrain DA system is composed of anatomically and functionally heterogeneous DA subpopulations with different axonal projections. These findings may explain a number of previously confusing observations that suggested a role for DA in processing both rewarding as well as aversive events. Here we will focus on recent advances in understanding the neural circuits mediating reward and aversion in the VTA and how stress as well as drugs of abuse, in particular cocaine, alter circuit function within a heterogeneous midbrain DA system. This article is part of a Special Issue entitled 'NIDA 40th Anniversary Issue'.

Optogenetic strategies to investigate neural circuitry engaged by stress.

Optogenetic techniques have given researchers unprecedented access to the function of discrete neural circuit elements and have been instrumental in the identification of novel brain pathways that become dysregulated in neuropsychiatric diseases. For example, stress is integrally linked to the manifestation and pathophysiology of neuropsychiatric illness, including anxiety, addiction and depression. Due to the heterogeneous populations of genetically and neurochemically distinct neurons in areas such as the bed nucleus of the stria terminalis (BNST), as well as their substantial number of projections, our understanding of how neural circuits become disturbed after stress has been limited. Using optogenetic tools, we are now able to selectively isolate distinct neural circuits that contribute to these disorders and perturb these circuits in vivo, which in turn may lead to the normalization of maladaptive behavior. This review will focus on current optogenetic strategies to identify, manipulate, and record from discrete neural circuit elements in vivo as well as highlight recent optogenetic studies that have been utilized to parcel out BNST function.