Severe and/or repeated stress exposure can lead to a number of maladaptive physiological and behavioral changes that contribute to psychiatric illnesses. Recent work indicates that the neuropeptide pituitary-adenylate-cyclase-activating-polypeptide (PACAP) plays an important role in stress-related psychopathologies relevant to depression and trauma-related disorders such as PTSD. However, the specific neural circuits that mediate PACAP effects on stress function are not fully understood. One candidate area is the lateral septum (LS), a limbic structure where PACAP and its cognate PAC1 receptors are abundantly expressed. Despite this neuroanatomical evidence, direct functional data supporting a role for septal PACAP/PAC1 receptor signaling in stress regulation are lacking. Using quantitative PCR, we show that forced swim stress increases PACAP mRNA expression in several limbic areas, including the LS, bed nucleus of the stria terminalis and basolateral amygdala, while chronic variable mild stress reduced PACAP expression in the LS only. Providing functional evidence of a PACAP/stress interaction, local administration of PACAP38 into the LS potentiated stress-induced ACTH release and altered stress-coping behavior by increasing passive (floating) and reducing active (struggling) coping during a forced swim challenge. Moreover, intraseptal PACAP38 administration significantly increased anxiety-like behavior in the elevated plus-maze and reduced grooming behavior in the sucrose splashtest, indicating anxiogenic and motivationally disruptive effects following enhanced PACAP signaling in the LS. Importantly, to assess the contribution of endogenous PACAP signaling, intra-LS administration of the PACAP receptor antagonist PACAP(6-38) produced a robust anxiolytic phenotype in the elevated plus-maze. Collectively, these findings provide the first direct evidence that PACAP/PAC1 receptor signaling in the LS modulates emotional and motivational processes in response to stress, identifying this system as a potential target for neuromodulatory interventions in stress-related psychiatric disorders.

Cancer-related cognitive impairment (CRCI) is increasingly recognized in patients with solid tumors and treated with immunotherapy but the mechanisms linking tumor immune status, systemic inflammation and neurocognitive dysfunction remain unclear. Here, we investigate in syngeneic immuno-desert (B16F10), immuno-excluded (B16F10 Ova) and immuno-inflamed (MC38) tumor-bearing immunocompetent male mice treated with anti-PD-1/anti-PD-L1 immune checkpoint inhibitors, behaviors, brain immune cells infiltration/homeostasis and neuroinflammation. Immuno-excluded and immuno-inflamed cancers impaired short-term memory as well as anxiety- and resignation-like behaviors, the anxiety-like behavior being correlated with an extended tumor “immunoscore”. Both immuno-desert and immuno-inflamed tumors evoked blood MCP-1/TNF- blood cytokines and meningeal STING-dependent transcriptional reprogramming with distinct B cell- and myeloid/T cell-dominated signatures. Immuno-inflamed MC38 tumors more specifically generated IL-6/IL-17 systemic inflammatory reaction associated with brain barriers permeability, ventriculomegaly, leptomeningeal myeloid cell accumulation, reactive hippocampal microglia and reduced neurogenesis. Anti-PD-1 and anti-PD-L1 did not affect behavior in cancer-naïve mice but exacerbated CRCI in a tumor-immune-status-dependent manner, increasing CD3+T cell infiltration, vascular inflammation, CCL19 expression and impacting progenitor proliferation in the dentate gyrus of the Hippocampus. Notably, anti-PD-L1 promoted BBB permeability, expanded circulating γδT cells and their accumulation at blood–brain barriers. γδT cell immunoneutralization prevented cognitive and anxiety-like deficits without impairing anti-PD-L1 efficacy. These findings identify barrier- and meningeal-centered mechanisms, and γδT cell-dependent pathways, as a central driver of immuno-inflamed- and PD-L1-related CRCI and provide new biomarkers and targets for neuroprotective strategies in cancer patients treated by immunotherapy.

Adolescence is a critical late phase of the neurodevelopment, characterized by marked brain plasticity and increased vulnerability to environmental challenges such as alcohol exposure. This study examined the impact of binge-like alcohol exposure in male Swiss Webster mice, focusing on oxidative damage, epigenetic and transcriptional alterations in key brain regions, such as the prefrontal cortex, cerebellum, striatum and hippocampus. Our results demonstrated that acute alcohol exposure during adolescence induces oxidative damage with significant alterations in global DNA methylation and gene expression involved in epigenetic regulation with distinct temporal and anatomical profiles. In the prefrontal cortex binge-like alcohol exposure exhibited persistent upregulation of genes associated with DNA methylation and histone deacetylation, consistent with prolonged transcriptional silencing that may impair executive functions and decision-making. The hippocampus appeared particularly sensitive, exhibiting marked decreases in DNA methylation and gene expression changes associated with an open chromatin state leading potentially linked to cognitive impairments in memory and learning impairments in memory and learning. In the striatum, binge-like alcohol exposure induced active DNA demethylation and transient modulation of histone methyltransferases, reflecting a dynamic compensatory response to alcohol-induced transcriptional repression, with implications for reward processing and impulse control. Similarly the cerebellum displayed a biphasic transcriptional pattern suggesting adaptive or homeostatic mechanisms aimed at maintaining cellular and synaptic balance. Collectively, these findings, accompanied by alterations in behavioral tests, highlight the regional specificity of epigenetic remodeling induced by excessive alcohol exposure during adolescence and offer new insights into the molecular mechanisms underlying increased neurodevelopmental vulnerability during this period.

Background/Objectives: Endozepines known as the endogenous ligands of benzodiazepine-binding sites, include the diazepam binding inhibitor (DBI) and its processing products, the triakontatetraneuropeptide (TTN) and the octadecaneuropeptide (ODN). Despite indisputable evidence of the binding of ODN on GABAAR-BZ-binding sites, their action on this receptor lacks compelling electrophysiological observations, with some studies reporting that ODN acts as a negative allosteric modulator (NAM) of GABAAR while others suggest the opposite (positive allosteric modulation, PAM effect). All these studies were carried out in vitro with various neuronal cell types. To further elucidate the role of ODN in neuronal excitability, we tested its effect in vivo in the cerebral cortex of the anesthetized mouse. Methods: Spontaneous neuronal spikes were recorded by means of an extracellular pipette, in the vicinity of which ODN was micro-infused, either at a high dose (10−5 M) or low dose (10−11 M). Results: ODN at a high dose induced a significant increase in neuronal spiking. This effect could be antagonized by the GABAAR-BZ-binding site blocker flumazenil. In sharp contrast, at low concentrations, ODN reduced neuronal spiking with a magnitude similar to GABA itself. Interestingly, this decrease in neuronal activity by low dose of ODN was not flumazenil-dependent, suggesting that this effect is mediated by another receptor. Finally, we show that astrocytes in culture, known to be stimulated by picomolar doses of ODN via a GPCR, increased their export of GABA when stimulated by low dose of ODN. Conclusion: Our results confirm the versatility of ODN in the control of GABA transmission, but suggest that its PAM-like effect is, at least in part, mediated via an astrocytic non-GABAAR ODN receptor release of GABA.

Spinal cord injury (SCI) leads to irreversible motor and sensory deficits and currently has no curative treatment. Among the various therapeutic strategies explored, cell transplantation— using either stem or differentiated cells—has been extensively studied. One of the most promising approaches involves the use of olfactory ensheathing cells (OECs), which have demonstrated unique potential to promote functional recovery and tissue repair following SCI. However, the mechanisms underlying these effects remain poorly understood. In this study, we investigated how OEC transplantation modulates endogenous spinal stem cells, with a particular focus on ependymal cells. Using inducible transgenic mouse lines and fate-mapping approaches, we show that OEC transplantation significantly enhances ependymal cell proliferation and self-renewal both in vivo and in vitro . Moreover, OECs promote astrocytic differentiation of ependymal progeny, which express low levels of inhibitory molecules— suggesting a supportive role in creating a permissive scar microenvironment. Transcriptomic analyses further revealed that OEC transplantation downregulates genes associated with axonal growth inhibition, thereby contributing to improved neuronal survival. Finally, by using a unique transgenic mouse model in which ependymal cell proliferation is genetically blocked, we demonstrate that the beneficial effects of OEC transplantation depend on ependymal cell activation. Together, these findings establish ependymal cells as essential mediators of the regenerative response induced by OECs. They also highlight a therapeutic strategy based on activating and modulating endogenous stem cells via the transient presence of non-integrating transplanted glial cells. This work contributes to our understanding of SCI repair and supports the clinical potential of OEC-based cell therapies. ### Competing Interest Statement The authors have declared no competing interest. ADIR Association, https://ror.org/01z9be204 Fondation de l'Avenir, https://ror.org/02dddmh90, AP-RMA-15-043 IRME association

Sterculia setigera is a medicinal plant of Togolese flora. We have previously reported that S. setigera leaves dry hydroethanolic extract (SSE) protects in vitro cerebellar granule neurons against H2O2 and 6-OHDA-induced cell death and in vivo against ethanol neurotoxicity on the cerebellar cortex of 8-day-old Wistar rats. The present study aimed to extend our knowledge of the protective effects of SSE by exploring its anti-apoptotic mechanisms on cultured PC12 cells. Apoptosis was induced by serum depletion before treatment with various concentrations (5-100 µg/mL) of SSE. SSE (20 µg/mL) significantly protected PC12 cells (+51.8 %) against serum depletion-induced PC12 cell death and inhibited Caspase-3/7 (-65.1 %) activity. SSE also significantly increased the expression of the anti-apoptotic factor Bcl-2. Conversely, it repressed the expression of Casp-3, Tp53 and Ddit3. Taken together, these results indicate that inhibition of apoptosis appears to be one of the main mechanisms of S. setigera neuroprotective effects.

Discovered in 1989, pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide with strong neuroprotective properties, as shown in various neurodegenerative preclinical models of Parkinson, Alzheimer, or Huntington diseases. PACAP neuroprotection has also been reported in animal models of cerebral ischemia and traumatic brain injury. The neuroprotective effect of PACAP occurs through its capacity to modulate most of the multiphasic aspects of neuronal diseases, such as oxidative stress, neuronal cell death, and inflammation. However, more than three decades after its discovery, and although PACAP neurotrophic and neuroprotective activities have now been largely documented, its clinical use is still awaited. Thus, the aim of this manuscript is to discuss the main reasons which limit the use of PACAP as a therapeutic agent for the treatment of neuronal diseases. To achieve this objective, an opinion survey has been conducted among experts in the field of PACAP, and a bibliographic investigation was carried out.

Migration is an essential characteristic of cells that occurs during many physiological and pathological processes. Astrocytes represent the most abundant cell type in the adult central nervous system (CNS), that play a crucial role in various functions such as guiding and supporting neuronal migration during development and maintaining brain homeostasis at adulthood. Astrocytes specifically synthesize and release endozepines, a family of regulatory peptides, including the octadecaneuropeptide (ODN). ODN is an endogenous ligand for both central- type benzodiazepine receptors and a metabotropic receptor. ODN promotes proliferation and prevents oxidative damage induced apoptosis on both neurons and astrocytes. However, little is known regarding the effect of ODN on cell migration. The purpose of the present study was to investigate the potential effect of ODN on astrocytes migration. Our results show that ODN stimulates astrocytes proliferation and migration at very low concentrations in wound healing assays, that was mimicked by the metabotropic ODN receptor agonist cyclo1-8 octapeptide (cyclo1-8OP, 10-14 M to 10-10 M). The effect of ODN on astrocyte migration was abrogated by the metabotropic receptor antagonist, cyclo1-8[DLeu5] OP. Moreover, we have shown that ODN activates the calcium signaling pathway and increases the mammalian target of rapamycin (mTOR) gene transcription, which are both known to promote astrocyte migration. Therefore, the present results suggest that ODN regulates astroglial cell migration through the calcium/mTOR signaling pathway and provide new insight regarding the role of ODN on brain remodling after injury.

Vasoactive intestinal peptide (VIP) is a highly abundant neuropeptide in the central nervous system, implicated in the regulation of numerous behavioral and physiological functions, including the central stress response. Notably, VIP and its cognate receptors, VPAC1 and VPAC2, are widely expressed in brain areas implicated in stress and anxiety regulation such as the amygdala. However, the exact role of VIP in stress function is not fully understood. The present study therefore examined how acute or chronic stress affect VIP and VPAC receptor gene expression in specific limbic brain regions of Sprague-Dawley rats using quantitative real-time PCR. Acute stress via forced swim exposure significantly increased VIP expression in the central amygdala (CeA) by 200 %, and in the medial amygdala (MeA) by 350 %. It also elevated VPAC1 receptor expression in the hypothalamic paraventricular nucleus (PVN), basolateral amygdala (BLA), and CeA, and VPAC2 receptor expression in the bed nucleus of the stria terminalis (BNST) and BLA. Chronic unpredictable stress induced both overlapping and distinct patterns, including upregulation of the VPAC1 receptor expression in the PVN by 120 % and the VPAC2 receptor in the CeA by 280 %, accompanied by slight downregulation of VIP in the CeA. These findings highlight a stress-duration-dependent and region-specific regulation of the VIP/VPAC receptor system and its potential role in modulating stress-related neural circuits.

The Concise Guide to Pharmacology 2025/26 marks the seventh edition in this series of biennial publications in the British Journal of Pharmacology. Presented in landscape format, the guide provides a comparative overview of the pharmacology of drug target families. The concise nature of the Concise Guide refers to the style of presentation, being clear, accessible, and well-structured, rather than the scope of the content, which spans approximately 500 pages. The Concise Guide summarises the key pharmacological properties of around 1900 human drug targets, and nearly 7000 interactions, involving around 4400 ligands. While the content is a substantially condensed version of the more detailed information and links available at the www.guidetopharmacology.org website, the printed guide serves as a permanent, citable, point-in-time record, that remains stable despite ongoing updates to the online database. The full contents of this publication can be found at https://bpspubs.onlinelibrary.wiley.com/doi/10.1111/bph.70230. The Concise Guides provide expert-curated recommendations of 'Gold Standard' selective pharmacological tools, available either commercially or as donations, which enable the identification of individual drug targets or families of drug targets. While the Concise Guide offers a more streamlined overview, more comprehensive information, including detailed pharmacological profiles and links to multiple online databases, is available through the Guide to Pharmacology website. The 2025/26 edition of the Concise Guide is based on material current as of mid-2025, and supersedes all previous editions, including the 2023/24 Guide, and earlier Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), and as such provides official IUPHAR classification and nomenclature for human drug targets, where applicable. G protein-coupled receptors are one of the six major pharmacological targets into which the Guide is divided, with the others being: ion channels, nuclear hormone receptors, catalytic receptors, enzymes and transporters. Each section includes nomenclature guidance, concise summaries, information of the best available pharmacological tools, key references, and suggestions for further reading.

Binge drinking (BD) is a widespread pattern of excessive alcohol consumption among adolescents and young adults with detrimental consequences for brain development. Animal models are essential for investigating the neurobiological mechanisms underlying BD, but selecting an appropriate model is critical to ensure relevance to human behavior. This study aims to validate a murine model of (BD) using Swiss Webster mice. To achieve this, both adolescent and adult mice were exposed to either a single binge (SB) or multiple binge (MB) of BD through intraperitoneal ethanol injections. The findings reveal that the SB protocol produces high blood alcohol concentrations (BACs) (150-400 mg/dL) sustained for several hours, with no significant differences based on age or episode repetition. However, the neurotoxic effects vary, showing that in adolescents, a single episode of BD reduces brain cell survival by 25 %, whereas in adults, multiple episodes are required to observe a 17 % decrease. This murine model of BD in Swiss Webster mice fulfills the main validation criteria identified in the literature. It presents valuable opportunities for studying individual variability and the neurobiological mechanisms of BD in adolescents, in order to identify potential therapeutic targets.

The cortex immediately surrounding a brain ischemic lesion, the peri-infarct cortex (PIC), harbors a large part of the potential to recover lost functions. However, our understanding of the neurophysiological conditions in which synaptic plasticity operates remains limited. Here we hypothesized that the chronic imbalance between excitation and inhibition of the PIC prevents the normalization of the gamma rhythm, a waveband of neural oscillations thought to orchestrate action potential trafficking. Probing the local field potential activity of the forelimb primary sensory cortex (S1FL) located in the PIC of male adult mice, we found a constant, deep reduction of low-gamma oscillation power (L-gamma; 30-50 Hz) precisely during the critical time window for recovery (1-3 weeks after stroke). The collapse of L-gamma power negatively correlated with behavioral progress in affected forelimb use. Mapping astrocyte reactivity and GABA-like immunoreactivity in the PIC revealed a parallel high signal, which gradually increased when approaching the lesion. Increasing tonic inhibition with local infusion of GABA or by blocking its recapture reduced L-gamma oscillation power in a magnitude similar to stroke. Conversely, the negative allosteric modulation of tonic GABA conductance using L655,708 or the gliopeptide ODN rescued the L-gamma power of the PIC. Altogether the present data point out that the chronic excess of ambient GABA in the PIC limits the generation of L-gamma oscillations in the repairing cortex and suggests that rehabilitative interventions aimed at normalizing low-gamma power within the critical period of stroke recovery could optimize the restitution of lost functions.

Zebra mussel (ZM), Dreissena polymorpha, commonly used as a sentinel species in freshwater biomonitoring, is now in competition for habitat with quagga mussel (QM), Dreissena rostriformis bugensis. This raises the question of the quagga mussel’s use in environmental survey. To better characterise QM response to stress compared with ZM, both species were exposed to cadmium (100 µg·L−1), a classic pollutant, for 7 days under controlled conditions. The gill proteomes were analysed using two-dimensional electrophoresis coupled with mass spectrometry. For ZM, 81 out of 88 proteoforms of variable abundance were identified using mass spectrometry, and for QM, 105 out of 134. Interestingly, the proteomic response amplitude varied drastically, with 5.6% of proteoforms of variable abundance (DAPs) in ZM versus 9.4% in QM. QM also exhibited greater cadmium accumulation. Only 12 common DAPs were observed. Several short proteoforms were detected, suggesting proteolysis. Functional analysis is consistent with the pleiotropic effects of the toxic metal ion cadmium, with alterations in sulphur and glutathione metabolisms, cellular calcium signalling, cytoskeletal dynamics, energy production, chaperone activation, and membrane events with numerous proteins involved in trafficking and endocytosis/exocytosis processes. Beyond common responses, the sister species display distinct reactions, with cellular response to stress being the main category involved in ZM as opposed to calcium and cytoskeleton alterations in QM. Moreover, QM exhibited greater evidence of proteolysis and cell death. Overall, these results suggest that QM has a weaker stress response capacity than ZM.

Astrocytes specifically synthesize and release endozepines, a family of regulatory peptides including octadecaneuropeptide (ODN). We have previously reported that ODN rescues neurons and astrocytes from 6-OHDA-induced oxidative stress and cell death. The purpose of this study was to examine the potential implication of miR-34b, miR-29a, and miR-21 in the protective activity of ODN on 6-OHDA-induced oxidative stress and cell death in cultured rat astrocytes. Flow cytometry analysis showed that 6-OHDA increased the number of early apoptotic and apoptotic dead cells while treatment with the subnanomolar dose of ODN significantly reduced the number of apoptotic cells induced by 6-OHDA. 6-OHDA-treated astrocytes exhibited the over-expression of miR-21 (+118%) associated with a knockdown of miR-34b (−61%) and miR-29a (−49%). Co-treatment of astrocytes with ODN blocked the 6-OHDA-stimulated production of ROS and NO and stimulation of Bax and caspase-3 gene transcription. Concomitantly, ODN down-regulated the expression of miR-34b and miR-29a and rescued the 6-OHDA-associated reduced expression of miR21, indicating that ODN regulates their expression during cell death. Transfection with miR-21-3p inhibitor prevented the effect of 6-OHDA against cell death. In conclusion, our study indicated that (i) the expression of miRNAs miR-34b, miR-29a, and miR-21 is modified in astrocytes under 6-OHDA injury and (ii) that ODN prevents this deregulation to induce its neuroprotective action. The present study identified miR-21 as an emerging candidate and as a promising pharmacological target that opens new neuroprotective therapeutic strategies in neurodegenerative diseases, especially in Parkinson’s disease.

Plants are a valuable source of information for pharmacological research and new drug discovery. The present study aimed to evaluate the neuroprotective potential of the leaves of the medicinal plant Sterculia setigera . In vitro , the effect of Sterculia setigera leaves dry hydroethanolic extract (SSE) was tested on cultured cerebellar granule neurons (CGN) survival when exposed to hydrogen peroxide (H 2 O 2 ) or 6-hydroxydopamine (6-OHDA), using the viability probe fluorescein diacetate (FDA), a lactate dehydrogenase (LDH) activity assay, an immunocytochemical staining against Gap 43, and quantification of the expression of genes involved in apoptosis, necrosis or oxidative stress. In vivo , the effect of intraperitoneal (ip) injection of SSE was assessed on the developing brain of 8-day-old Wistar rats exposed to ethanol neurotoxicity by measuring caspase-3 activity on cerebellum homogenates, the expression of some genes in tissue extracts, the thickness of cerebellar cortical layers and motor coordination. In vitro , SSE protected CGN against H 2 O 2 and 6-OHDA induced cell death at a dose of 10 µg/mL, inhibited the expression of genes Casp3 and Bad , and upregulated the expression of Cat and Gpx7 . In vivo , SSE significantly blocked the deleterious effect of ethanol by reducing the activity of caspase-3, inhibiting the expression of Bax and Tp53 , preventing the reduction of the thickness of the internal granule cell layer of the cerebellar cortex and restoring motor functions. Sterculia setigera exerts neuroactive functions as claimed by traditional medicine and should be a good candidate for the development of a neuroprotective treatment against neurodegenerative diseases.

The Concise Guide to PHARMACOLOGY 2023/24 is the sixth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of approximately 1800 drug targets, and about 6000 interactions with about 3900 ligands. There is an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes almost 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at . G protein-coupled receptors are one of the six major pharmacological targets into which the Guide is divided, with the others being: ion channels, nuclear hormone receptors, catalytic receptors, enzymes and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2023, and supersedes data presented in the 2021/22, 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.

Apnea of prematurity (AOP) affects more than 50% of preterm infants and leads to perinatal intermittent hypoxia (IH) which is a major cause of morbimortality worldwide. At birth, the human cerebellar cortex is still immature, making it vulnerable to perinatal events. Additionally, studies have shown a correlation between cerebellar functions and the deficits observed in children who have experienced AOP. Yet, the cerebellar alterations underpinning this link remain poorly understood. To gain insight into the involvement of the cerebellum in perinatal hypoxia-related consequences, we developed a mouse model of AOP. Our previous research has revealed that IH induces oxidative stress in the developing cerebellum, as evidenced by the over-expression of genes involved in reactive oxygen species production and the under-expression of genes encoding antioxidant enzymes. These changes suggest a failure of the defense system against oxidative stress and could be responsible for neuronal death in the cerebellum.Building upon these findings, we conducted a transcriptomic study of the genes involved in the processes that occur during cerebellar development. Using real-time PCR, we analyzed the expression of these genes at different developmental stages and in various cell types. This enabled us to pinpoint a timeframe of vulnerability at P8, which represents the age with the highest number of downregulated genes in the cerebellum. Furthermore, we discovered that our IH protocol affects several molecular pathways, including proliferation, migration, and differentiation. This indicates that IH can impact the development of different cell types, potentially contributing to the histological and behavioral deficits observed in this model. Overall, our data strongly suggest that the cerebellum is highly sensitive to IH, and provide valuable insights into the cellular and molecular mechanisms underlying AOP. In the long term, these findings may contribute to the identification of novel therapeutic targets for improving the clinical management of this prevalent pathology.

Background: PC12 pheochromocytoma tumor cell lines are widely used to decipher the intracellular signaling mechanisms mediating the effects of some growth factors. Nevertheless, the disparity in appearance of some PC12 cell lines used in the different publications questions our ability to compare the results obtained by the numerous laboratories which use them. This led us to analyze the phenotypic aspect and transcriptomic expression of 5 PC12 cell lines from different origins under control conditions and after treatment with nerve growth factor (NGF) or pituitary adenylate cyclase-activating polypeptide (PACAP). Methods: Characterization of the 5 PC12 cell lines was conducted using imaging techniques and high-throughput real-time PCR combined with bioinformatics analysis. Results: The results show that the 5 cell lines are very variable in terms of shape, proliferation rate, motility, adhesion to the substrate, and gene expression. This high heterogeneity of the cell lines is also found when looking at their response to NGF or PACAP on gene expression or differentiation, with even in some cases opposite effects, as, for example, on cell proliferation. Actually, only 2 of the cell lines tested exhibited some phenotypic similarities with each other, even though the transcriptomic analyses show that they are far from identical. Discussion/Conclusion: As this issue of cell heterogenicity is not restricted to PC12 cells, the present results highlight the need to facilitate the supply of cell lines at low cost, the necessity to standardize practices regarding the use of cell lines, and the requirement to define precise markers of established cell lines which should be monitored in every publication. Regarding this latter point, the present data show that transcriptomic analysis by real-time PCR using a panel of genes of interest is easy to implement and provides a reliable method to control the possible drift of the cells over time in culture. Transcriptomic phenotyping combined with bioinformatics analysis can also be a useful approach to predict the response of the cells to treatments in terms of cell signaling activation, which can help to choose among several cell lines the most appropriate one for the investigation of a particular mechanism. Taken together, the results from this study highlight the need to use well-characterized cell lines with standardized protocols to generate reproducible results from 1 laboratory to the other.