Impact of light-emitting diodes on visual cortex layer 5 pyramidal neurons (V1-L5PNs)-A rodent study.

Purpose: Light-induced neural retinal insult leads to alterations in the visual cortex neurons. We examined light-induced neuronal alterations in the visual cortex layer 5 pyramidal neurons (V1-L5PNs) of adult male Wistar rats. Methods: A total of 24 rats were divided into the following groups (n=6 each): control (NC), blue (BL), white (WL), and yellow (YL). The exposure groups were subjected to light-emitting diodes (LED) exposure (450-500 lx) of differing wavelengths for 90 days (12:12 16 light-dark cycle). After LED exposure, the animals were sacrificed, and the brain tissues were removed and impregnated in freshly prepared Golgi-Cox stain for 21 days. Sholl's grading analysis was used to quantify the apical and basal dendritic branching points and intersections of the V1-L5PNs. Results: There was a significant difference in the number of apical branching points among all groups (p<0.001), with a particularly notable difference between the BL and WL groups (p<0.001). A post hoc test revealed that all exposure groups (BL, WL, and YL) had fewer apical branching points (p<0.001) on an average of 3.6 µm and a significant reduction in the dendritic intersections (p<0.001) compared to the number of branching points extending from layer Va (V1) neurons. Conclusions: Chronic and cumulative exposure to blue and white LEDs led to the pruning of V1-L5PNs, which might impair visual processing.

Functional specialization of hippocampal somatostatin-expressing interneurons.

Hippocampal somatostatin-expressing (Sst) GABAergic interneurons (INs) exhibit considerable anatomical and functional heterogeneity. Recent single-cell transcriptome analyses have provided a comprehensive Sst-IN subpopulations census, a plausible molecular ground truth of neuronal identity whose links to specific functionality remain incomplete. Here, we designed an approach to identify and access subpopulations of Sst-INs based on transcriptomic features. Four mouse models based on single or combinatorial Cre- and Flp- expression differentiated functionally distinct subpopulations of CA1 hippocampal Sst-INs that largely tiled the morpho-functional parameter space of the Sst-INs superfamily. Notably, the Sst;;Tac1 intersection revealed a population of bistratified INs that preferentially synapsed onto fast-spiking interneurons (FS-INs) and were sufficient to interrupt their firing. In contrast, the Ndnf;;Nkx2-1 intersection identified a population of oriens lacunosum-moleculare INs that predominantly targeted CA1 pyramidal neurons, avoiding FS-INs. Overall, our results provide a framework to translate neuronal transcriptomic identity into discrete functional subtypes that capture the diverse specializations of hippocampal Sst-INs.

Directed and acyclic synaptic connectivity in the human layer 2-3 cortical microcircuit.

The computational capabilities of neuronal networks are fundamentally constrained by their specific connectivity. Previous studies of cortical connectivity have mostly been carried out in rodents; whether the principles established therein also apply to the evolutionarily expanded human cortex is unclear. We studied network properties within the human temporal cortex using samples obtained from brain surgery. We analyzed multineuron patch-clamp recordings in layer 2-3 pyramidal neurons and identified substantial differences compared with rodents. Reciprocity showed random distribution, synaptic strength was independent from connection probability, and connectivity of the supragranular temporal cortex followed a directed and mostly acyclic graph topology. Application of these principles in neuronal models increased dimensionality of network dynamics, suggesting a critical role for cortical computation.

Anterior cingulate cortex and insomnia: A cingulate-striatum connection.

Insomnia is an important comorbidity of chronic pain. In this issue of Neuron, Li et al. report that chronic-pain-induced insomnia is mediated by the pyramidal neurons in the anterior cingulate cortex and their dopaminergic projections to the dorsal medial striatum.

Morphological correlates of pyramidal cell axonal myelination in mouse and human neocortex.

The axons of neocortical pyramidal neurons are frequently myelinated. Heterogeneity in the topography of axonal myelination in the cerebral cortex has been attributed to a combination of electrophysiological activity, axonal morphology, and neuronal-glial interactions. Previously, we showed that axonal segment length and caliber are critical local determinants of fast-spiking interneuron myelination. However, the factors that determine the myelination of individual axonal segments along neocortical pyramidal neurons remain largely unexplored. Here, we used structured illumination microscopy to examine the extent to which axonal morphology is predictive of the topography of myelination along neocortical pyramidal neurons. We identified critical thresholds for axonal caliber and interbranch distance that are necessary, but not sufficient, for myelination of pyramidal cell axons in mouse primary somatosensory cortex (S1). Specifically, we found that pyramidal neuron axonal segments with a caliber < 0.24 µm or interbranch distance < 18.10 µm are rarely myelinated. Moreover, we further confirmed that these findings in mice are similar for human neocortical pyramidal cell myelination (caliber < 0.25 µm, interbranch distance < 19.00 µm), suggesting that this mechanism is evolutionarily conserved. Taken together, our findings suggest that axonal morphology is a critical correlate of the topography and cell-type specificity of neocortical myelination.

Agranular frontal cortical microcircuit underlying cognitive control in macaques.

The error-related negativity and an N2-component recorded over medial frontal cortex index core functions of cognitive control. While they are known to originate from agranular frontal areas, the underlying microcircuit mechanisms remain elusive. Most insights about microcircuit function have been derived from variations of the so-called canonical microcircuit model. These microcircuit architectures are based extensively on studies from granular sensory cortical areas in monkeys, cats, and rodents. However, evidence has shown striking cytoarchitectonic differences across species and differences in the functional relationships across cortical layers in agranular compared to granular sensory areas. In this minireview, we outline a tentative microcircuit model underlying cognitive control in the agranular frontal cortex of primates. The model incorporates the main GABAergic interneuron subclasses with specific laminar arrangements and target regions on pyramidal cells. We emphasize the role of layer 5 pyramidal cells in error and conflict detection. We offer several specific questions necessary for creating a specific intrinsic microcircuit model of the agranular frontal cortex.

Alfaxalone is an effective anesthetic for the electrophysiological study of anoxia-tolerance mechanisms in western painted turtle pyramidal neurons.

Anoxia in the mammalian brain leads to hyper-excitability and cell death; however, this cascade of events does not occur in the anoxia-tolerant brain of the western painted turtle, Chrysemys picta belli. The painted turtle has become an important anoxia-tolerant model to study brain, heart, and liver function in the absence of oxygen, but being anoxia-tolerant likely means that decapitation alone is not a suitable method of euthanasia. Many anesthetics have long-term effects on ion channels and are not appropriate for same day experimentation. Using whole-cell electrophysiological techniques, we examine the effects of the anesthetic, Alfaxalone, on pyramidal cell action potential amplitude, threshold, rise and decay time, width, frequency, whole cell conductance, and evoked GABAA receptors currents to determine if any of these characteristics are altered with the use of Alfaxalone for animal sedation. We find that Alfaxalone has no long-term impact on action potential parameters or whole-cell conductance. When acutely applied to naïve tissue, Alfaxalone did lengthen GABAA receptor current decay rates by 1.5-fold. Following whole-animal sedation with Alfaxalone, evoked whole cell GABAA receptor current decay rates displayed an increasing trend with 1 and 2 hours after brain sheet preparation, but showed no significant change after a 3-hour washout period. Therefore, we conclude that Alfaxalone is a suitable anesthetic for same day use in electrophysiological studies in western painted turtle brain tissue.

Enhanced Release Probability without Changes in Synaptic Delay during Analogue-Digital Facilitation.

Neuronal timing with millisecond precision is critical for many brain functions such as sensory perception, learning and memory formation. At the level of the chemical synapse, the synaptic delay is determined by the presynaptic release probability (Pr) and the waveform of the presynaptic action potential (AP). For instance, paired-pulse facilitation or presynaptic long-term potentiation are associated with reductions in the synaptic delay, whereas paired-pulse depression or presynaptic long-term depression are associated with an increased synaptic delay. Parallelly, the AP broadening that results from the inactivation of voltage gated potassium (Kv) channels responsible for the repolarization phase of the AP delays the synaptic response, and the inactivation of sodium (Nav) channels by voltage reduces the synaptic latency. However, whether synaptic delay is modulated during depolarization-induced analogue-digital facilitation (d-ADF), a form of context-dependent synaptic facilitation induced by prolonged depolarization of the presynaptic neuron and mediated by the voltage-inactivation of presynaptic Kv1 channels, remains unclear. We show here that despite Pr being elevated during d-ADF at pyramidal L5-L5 cell synapses, the synaptic delay is surprisingly unchanged. This finding suggests that both Pr- and AP-dependent changes in synaptic delay compensate for each other during d-ADF. We conclude that, in contrast to other short- or long-term modulations of presynaptic release, synaptic timing is not affected during d-ADF because of the opposite interaction of Pr- and AP-dependent modulations of synaptic delay.

Different states of synaptic vesicle priming explain target cell type-dependent differences in neurotransmitter release.

Pronounced differences in neurotransmitter release from a given presynaptic neuron, depending on the synaptic target, are among the most intriguing features of cortical networks. Hippocampal pyramidal cells (PCs) release glutamate with low probability to somatostatin expressing oriens-lacunosum-moleculare (O-LM) interneurons (INs), and the postsynaptic responses show robust short-term facilitation, whereas the release from the same presynaptic axons onto fast-spiking INs (FSINs) is ~10-fold higher and the excitatory postsynaptic currents (EPSCs) display depression. The mechanisms underlying these vastly different synaptic behaviors have not been conclusively identified. Here, we applied a combined functional, pharmacological, and modeling approach to address whether the main difference lies in the action potential-evoked fusion or else in upstream priming processes of synaptic vesicles (SVs). A sequential two-step SV priming model was fitted to the peak amplitudes of unitary EPSCs recorded in response to complex trains of presynaptic stimuli in acute hippocampal slices of adult mice. At PC-FSIN connections, the fusion probability (Pfusion) of well-primed SVs is 0.6, and 44% of docked SVs are in a fusion-competent state. At PC-O-LM synapses, Pfusion is only 40% lower (0.36), whereas the fraction of well-primed SVs is 6.5-fold smaller. Pharmacological enhancement of fusion by 4-AP and priming by PDBU was recaptured by the model with a selective increase of Pfusion and the fraction of well-primed SVs, respectively. Our results demonstrate that the low fidelity of transmission at PC-O-LM synapses can be explained by a low occupancy of the release sites by well-primed SVs.

Electrophysiological and morphological properties of prefrontal pyramidal neurons innervated by mediodorsal thalamus.

The high-order cognitive and executive functions are necessary for an individual to survive. The densely bidirectional innervations between the medial prefrontal cortex (mPFC) and the mediodorsal thalamus (MD) play a vital role in regulating high-order functions. Pyramidal neurons in mPFC have been classified into several subclasses according to their morphological and electrophysiological properties, but the properties of the input-specific pyramidal neurons in mPFC remain poorly understood. The present study aimed to profile the morphological and electrophysiological properties of mPFC pyramidal neurons innervated by MD. In the past, the studies for characterizing the morphological and electrophysiological properties of neurons mainly relied on the electrophysiological recording of a large number of neurons and their morphologic reconstructions. But, it is a low efficient method for characterizing the circuit-specific neurons. The present study combined the advantages of traditional morphological and electrophysiological methods with machine learning to address the shortcomings of the past method, to establish a classification model for the morphological and electrophysiological properties of mPFC pyramidal neurons, and to achieve more accurate and efficient identification of the properties from a small size sample of neurons. We labeled MD-innervated pyramidal neurons of mPFC using the trans-synaptic neural circuitry tracing method and obtained their morphological properties using whole-cell patch-clamp recording and morphologic reconstructions. The results showed that the classification model established in the present study could predict the electrophysiological properties of MD-innervated pyramidal neurons based on their morphology. MD-innervated pyramidal neurons exhibit larger basal dendritic length but lower apical dendrite complexity compared to non-MD-innervated neurons in the mPFC. The morphological characteristics of the two subtypes (ET-1 and ET-2) of mPFC pyramidal neurons innervated by MD are different, with the apical dendrites of ET-1 neurons being longer and more complex than those of ET-2 neurons. These results suggest that the electrophysiological properties of MD- innervated pyramidal neurons within mPFC correlate with their morphological properties, indicating that the different roles of these two subclasses in local circuits within PFC, as well as in PFC-cortical/subcortical brain region circuits.

Inferring Parameters of Pyramidal Neuron Excitability in Mouse Models of Alzheimer's Disease Using Biophysical Modeling and Deep Learning.

Alzheimer's disease (AD) is believed to occur when abnormal amounts of the proteins amyloid beta and tau aggregate in the brain, resulting in a progressive loss of neuronal function. Hippocampal neurons in transgenic mice with amyloidopathy or tauopathy exhibit altered intrinsic excitability properties. We used deep hybrid modeling (DeepHM), a recently developed parameter inference technique that combines deep learning with biophysical modeling, to map experimental data recorded from hippocampal CA1 neurons in transgenic AD mice and age-matched wildtype littermate controls to the parameter space of a conductance-based CA1 model. Although mechanistic modeling and machine learning methods are by themselves powerful tools for approximating biological systems and making accurate predictions from data, when used in isolation these approaches suffer from distinct shortcomings: model and parameter uncertainty limit mechanistic modeling, whereas machine learning methods disregard the underlying biophysical mechanisms. DeepHM addresses these shortcomings by using conditional generative adversarial networks to provide an inverse mapping of data to mechanistic models that identifies the distributions of mechanistic modeling parameters coherent to the data. Here, we demonstrated that DeepHM accurately infers parameter distributions of the conductance-based model on several test cases using synthetic data generated with complex underlying parameter structures. We then used DeepHM to estimate parameter distributions corresponding to the experimental data and infer which ion channels are altered in the Alzheimer's mouse models compared to their wildtype controls at 12 and 24 months. We found that the conductances most disrupted by tauopathy, amyloidopathy, and aging are delayed rectifier potassium, transient sodium, and hyperpolarization-activated potassium, respectively.

Silencing CA1 pyramidal cells output reveals the role of feedback inhibition in hippocampal oscillations.

The precise temporal coordination of neural activity is crucial for brain function. In the hippocampus, this precision is reflected in the oscillatory rhythms observed in CA1. While it is known that a balance between excitatory and inhibitory activity is necessary to generate and maintain these oscillations, the differential contribution of feedforward and feedback inhibition remains ambiguous. Here we use conditional genetics to chronically silence CA1 pyramidal cell transmission, ablating the ability of these neurons to recruit feedback inhibition in the local circuit, while recording physiological activity in mice. We find that this intervention leads to local pathophysiological events, with ripple amplitude and intrinsic frequency becoming significantly larger and spatially triggered local population spikes locked to the trough of the theta oscillation appearing during movement. These phenotypes demonstrate that feedback inhibition is crucial in maintaining local sparsity of activation and reveal the key role of lateral inhibition in CA1 in shaping circuit function.

Increased NLRP1 mRNA and Protein Expression Suggests Inflammasome Activation in the Dorsolateral Prefrontal and Medial Orbitofrontal Cortex in Schizophrenia.

Schizophrenia is a complex mental condition, with key symptoms marked for diagnosis including delusions, hallucinations, disorganized thinking, reduced emotional expression, and social dysfunction. In the context of major developmental hypotheses of schizophrenia, notably those concerning maternal immune activation and neuroinflammation, we studied NLRP1 expression and content in the postmortem brain tissue of 10 schizophrenia and 10 control subjects. In the medial orbitofrontal cortex (Brodmann's area 11/12) and dorsolateral prefrontal cortex (area 46) from both hemispheres of six schizophrenia subjects, the NLRP1 mRNA expression was significantly higher than in six control brains (p < 0.05). As the expression difference was highest for the medial orbitofrontal cortex in the right hemisphere, we assessed NLRP1-immunoreactive pyramidal neurons in layers III, V, and VI in the medial orbitofrontal cortex in the right hemisphere of seven schizophrenia and five control brains. Compared to controls, we quantified a significantly higher number of NLRP1-positive pyramidal neurons in the schizophrenia brains (p < 0.01), suggesting NLRP1 inflammasome activation in schizophrenia subjects. Layer III pyramidal neuron dysfunction aligns with working memory deficits, while impairments of pyramidal neurons in layers V and VI likely disrupt predictive processing. We propose NLRP1 inflammasome as a potential biomarker and therapeutic target in schizophrenia.

Hippocampal Pyk2 regulates specific social skills: Implications for schizophrenia.

Pyk2 has been shown previously to be involved in several psychological and cognitive alterations related to stress, Huntington's disease, and Alzheimer's disease. All these disorders are accompanied by different types of impairments in sociability, which has recently been linked to improper mitochondrial function. We hypothesize that Pyk2, which regulates mitochondria, could be associated with the regulation of mitochondrial dynamics and social skills. In the present manuscript, we report that a reduction of Pyk2 levels in mouse pyramidal neurons of the hippocampus decreased social dominance and aggressivity. Furthermore, social interactions induced robust Pyk2-dependent hippocampal changes in several oxidative phosphorylation complexes. We also observed that Pyk2 levels were increased in the CA1 pyramidal neurons of schizophrenic subjects, occurring alongside changes in different direct and indirect regulators of mitochondrial function including DISC1 and Grp75. Accordingly, overexpressing Pyk2 in hippocampal CA1 pyramidal cells mimicked some specific schizophrenia-like social behaviors in mice. In summary, our results indicate that Pyk2 might play a role in regulating specific social skills likely via mitochondrial dynamics and that there might be a link between Pyk2 levels in hippocampal neurons and social disturbances in schizophrenia.

Low-intensity transcranial ultrasound stimulation improves memory in vascular dementia by enhancing neuronal activity and promoting spine formation.

Memory is closely associated with neuronal activity and dendritic spine formation. Low-intensity transcranial ultrasound stimulation (TUS) improves the memory of individuals with vascular dementia (VD). However, it is unclear whether neuronal activity and dendritic spine formation under ultrasound stimulation are involved in memory improvement in VD. In this study, we found that seven days of TUS improved memory in VD model while simultaneously increasing pyramidal neuron activity, promoting dendritic spine formation, and reducing dendritic spine elimination. These effects lasted for 7 days but disappeared on 14 d after TUS. Neuronal activity and dendritic spine formation strongly corresponded to improvements in memory behavior over time. In addition, we also found that the memory, neuronal activity and dendritic spine of VD mice cannot be restored again by TUS of 7 days after 28 d. Collectively, these findings suggest that TUS increases neuronal activity and promotes dendritic spine formation and is thus important for improving memory in patients with VD.

Plasma membrane calcium ATPase powered by glycolysis is the main mechanism for calcium clearance in the hippocampal pyramidal neuron.

AIMS: This study sought to elucidate the primary ATP-dependent mechanisms involved in clearing cytosolic Ca2+ in neurons and determine the predominant ATP-generating pathway-glycolysis or tricarboxylic acid cycle/oxidative phosphorylation (TCA/OxPhos)-associated with these mechanisms in hippocampal pyramidal neurons. MAIN METHODS: Our investigation involved evaluating basal Ca2+ levels and analyzing the kinetic characteristics of evoked neuronal Ca2+ transients after selectively combined the inhibition/blockade of key ATP-dependent mechanisms with the suppression of either TCA/OxPhos or glycolytic ATP sources. KEY FINDINGS: Our findings unveiled that the plasma membrane Ca2+ ATPase (PMCA) serves as the principal ATP-dependent mechanism for clearance cytosolic Ca2+ in hippocampal pyramidal neurons, both during rest and neuronal activity. Remarkably, during cellular activity, PMCA relies on ATP derived from glycolysis, challenging the traditional notion of neuronal reliance on TCA/OxPhos for ATP. Other mechanisms for Ca2+ clearance in pyramidal neurons, such as SERCA and NCX, appear to be dependent on TCA/OxPhos. Interestingly, at rest, the ATP required to fuel PMCA and SERCA, the two main mechanisms to keep resting Ca2+, seems to originate from a source other than glycolysis or the TCA/OxPhos. SIGNIFICANCE: These findings underscore the vital role of glycolysis in bolstering PMCA neuronal function to uphold Ca2+ homeostasis. Moreover, they elucidate the varying dependencies of cytoplasmic Ca2+ clearance mechanisms on distinct energy sources for their operation.

Prelimbic cortical pyramidal neurons to ventral tegmental area projections promotes arousal from sevoflurane anesthesia.

AIMS: General anesthesia has been used in surgical procedures for approximately 180 years, yet the precise mechanism of anesthetic drugs remains elusive. There is significant anatomical connectivity between the ventral tegmental area (VTA) and the prelimbic cortex (PrL). Projections from VTA dopaminergic neurons (VTADA ) to the PrL play a role in the transition from sevoflurane anesthesia to arousal. It is still uncertain whether the prelimbic cortex pyramidal neuron (PrLPyr ) and its projections to VTA (PrLPyr -VTA) are involved in anesthesia-arousal regulation. METHODS: We employed chemogenetics and optogenetics to selectively manipulate neuronal activity in the PrLPyr -VTA pathway. Electroencephalography spectra and burst-suppression ratios (BSR) were used to assess the depth of anesthesia. Furthermore, the loss or recovery of the righting reflex was monitored to indicate the induction or emergence time of general anesthesia. To elucidate the receptor mechanisms in the PrLPyr -VTA projection's impact on anesthesia and arousal, we microinjected NMDA receptor antagonists (MK-801) or AMPA receptor antagonists (NBQX) into the VTA. RESULTS: Our findings show that chemogenetic or optogenetic activation of PrLPyr neurons prolonged anesthesia induction and promoted emergence. Additionally, chemogenetic activation of the PrLPyr -VTA neural pathway delayed anesthesia induction and promoted anesthesia emergence. Likewise, optogenetic activation of the PrLPyr -VTA projections extended the induction time and facilitated emergence from sevoflurane anesthesia. Moreover, antagonizing NMDA receptors in the VTA attenuates the delayed anesthesia induction and promotes emergence caused by activating the PrLPyr -VTA projections. CONCLUSION: This study demonstrates that PrLPyr neurons and their projections to the VTA are involved in facilitating emergence from sevoflurane anesthesia, with the PrLPyr -VTA pathway exerting its effects through the activation of NMDA receptors within the VTA.

Anterior cingulate cross-hemispheric inhibition via the claustrum resolves painful sensory conflict.

The anterior cingulate cortex (ACC) responds to noxious and innocuous sensory inputs, and integrates them to coordinate appropriate behavioral reactions. However, the role of the projections of ACC neurons to subcortical areas and their influence on sensory processing are not fully investigated. Here, we identified that ACC neurons projecting to the contralateral claustrum (ACC→contraCLA) preferentially respond to contralateral mechanical sensory stimulation. These sensory responses were enhanced during attending behavior. Optogenetic activation of ACC→contraCLA neurons silenced pyramidal neurons in the contralateral ACC by recruiting local circuit fast-spiking interneuron activation via an excitatory relay in the CLA. This circuit activation suppressed withdrawal behavior to mechanical stimuli ipsilateral to the ACC→contraCLA neurons. Chemogenetic silencing showed that the cross-hemispheric circuit has an important role in the suppression of contralateral nociceptive behavior during sensory-driven attending behavior. Our findings identify a cross-hemispheric cortical-subcortical-cortical arc allowing the brain to give attentional priority to competing innocuous and noxious inputs.

Emergence of co-tuning in inhibitory neurons as a network phenomenon mediated by randomness, correlations, and homeostatic plasticity.

Cortical excitatory neurons show clear tuning to stimulus features, but the tuning properties of inhibitory interneurons are ambiguous. While inhibitory neurons have been considered to be largely untuned, some studies show that some parvalbumin-expressing (PV) neurons do show feature selectivity and participate in co-tuned subnetworks with pyramidal neurons. In this study, we first use mean-field theory to demonstrate that a combination of homeostatic plasticity governing the synaptic dynamics of the connections from PV to excitatory neurons, heterogeneity in the excitatory postsynaptic potentials that impinge on PV neurons, and shared correlated input from layer 4 results in the functional and structural self-organization of PV subnetworks. Second, we show that structural and functional feature tuning of PV neurons emerges more clearly at the network level, i.e., that population-level measures identify functional and structural co-tuning of PV neurons that are not evident in pairwise individual-level measures. Finally, we show that such co-tuning can enhance network stability at the cost of reduced feature selectivity.

Adenosinergic Modulation of Layer 6 Microcircuitry in the Medial Prefrontal Cortex Is Specific to Presynaptic Cell Type.

Adenosinergic modulation in the PFC is recognized for its involvement in various behavioral aspects including sleep homoeostasis, decision-making, spatial working memory and anxiety. While the principal cells of layer 6 (L6) exhibit a significant morphological diversity, the detailed cell-specific regulatory mechanisms of adenosine in L6 remain unexplored. Here, we quantitatively analyzed the morphological and electrophysiological parameters of L6 neurons in the rat medial prefrontal cortex (mPFC) using whole-cell recordings combined with morphological reconstructions. We were able to identify two different morphological categories of excitatory neurons in the mPFC of both juvenile and young adult rats with both sexes. These categories were characterized by a leading dendrite that was oriented either upright (toward the pial surface) or inverted (toward the white matter). These two excitatory neuron subtypes exhibited different electrophysiological and synaptic properties. Adenosine at a concentration of 30 µM indiscriminately suppressed connections with either an upright or an inverted presynaptic excitatory neuron. However, using lower concentrations of adenosine (10 µM) revealed that synapses originating from L6 upright neurons have a higher sensitivity to adenosine-induced inhibition of synaptic release. Adenosine receptor activation causes a reduction in the probability of presynaptic neurotransmitter release that could be abolished by specifically blocking A1 adenosine receptors (A1ARs) using 8-cyclopentyltheophylline (CPT). Our results demonstrate a differential expression level of A1ARs at presynaptic sites of two functionally and morphologically distinct subpopulations of L6 principal neurons, suggesting the intricate functional role of adenosine in neuronal signaling in the brain.