About

Ezequiel Marron Fernandez de Velasco, PhD, is an Associate Professor in the Department of Pharmacology at the University of Minnesota. He is also the Manager of the Viral Vector and Cloning Core and the Viral Innovation Core at the university. Dr. Marron received his B.S. in Biology and B.S. in Biochemistry, as well as M.S. degrees in Neuroscience and Education from the University of Salamanca. He earned his PhD in Biology/Neuroscience working in the laboratory of Dr. Raquel E. Rodriguez. After completing postdoctoral training in the laboratory of Dr. David Armstrong at NIEHS–NIH, he joined Dr. Kevin Wickman's laboratory at the University of Minnesota, where he worked as a Senior Scientist until his promotion in 2019.

Research topics

• Biology
• Neuroscience
• Medicine
• Internal medicine
• Pharmacology
• Cell biology
• Chemistry
• Genetics
• Physics
• Psychology
• Biophysics
• Developmental psychology

Selected publications

• Modulation of NMDA Receptor and TRPM4 Activity in Hippocampal Neurons with the NMDA Receptor/TRPM4 Interface Inhibitor Brophenexin

  Neurotoxicity Research · 2026-03-06

  articleOpen access

  Excitotoxic signaling mediated by N-methyl-D-aspartate receptors (NMDARs) is inhibited by NMDAR/TRPM4 complex inhibitors such as brophenexin (BPN). We used rat hippocampal neurons grown in culture to determine the effects of BPN on NMDAR and TRPM4 function. NMDA evoked concentration-dependent increases in intracellular Ca2+ that were inhibited by 10 µM BPN in a non-competitive manner. In contrast, the TRPM4 inhibitor 4-chloro-2-(2-(naphthalene-1-yloxy) acetamido) benzoic acid (NBA) increased the potency of NMDA at 22 °C. BPN inhibition of NMDAR-mediated increases in Ca2+ was fully reversible and recovered by rapid (30 s) and slow (90 min) processes. The rapid phase of recovery from BPN inhibition was mediated by trafficking through recycling endosomes as indicated by blockade of this phase with bafilomycin A1 an agent that prevents endosomal acidification. The full recovery of NMDAR function observed 90 min after washout of BPN was not affected by this treatment. Immunocytochemistry experiments suggested that BPN did not directly alter NMDAR trafficking but instead changes in surface and internal GluN2B immunoreactivity were likely homeostatic responses to inhibition of NMDAR function by BPN. In the presence of MK-801 to block NMDARs, 10 µM NBA inhibited spontaneous network-driven Ca2+ spiking by 74 ± 11% whereas 10 µM BPN reduced activity by 49 ± 6%. Thus, BPN inhibits TRPM4-dependent activity. In summary, BPN is a non-competitive, fully reversible inhibitor of NMDAR-mediated Ca2+ influx and produces a modest inhibition of TRPM4 function. Functional inhibition of these ion channels likely contributes to the neuroprotective properties of NMDAR/TRPM4 interface inhibitors.

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• AAV-only targeting of ventral tegmental area dopamine neurons for optical self-stimulation studies in mice

  bioRxiv (Cold Spring Harbor Laboratory) · 2026-02-12

  articleOpen access1st authorCorresponding

  Studies employing optogenetic approaches in rodent models have highlighted the important contribution of ventral tegmental area (VTA) dopamine (DA) neurons to reward, learning, and motivation. Selective manipulation of VTA DA neurons is generally achieved in these studies using transgenic mouse or rat lines that express Cre recombinase under the control of a promoter active in DA neurons, combined with intra-VTA infusion of adeno-associated virus (AAV) vectors harboring Cre recombinase-dependent expression cassettes. Reliance on transgenic Cre driver lines is expensive and decreases study efficiency, and available driver lines have unique limitations. Here, we report the development of an AAV-only approach that permits genetic access to VTA DA neurons and can support optogenetic self-stimulation in mice. We used a 2.5 kb fragment of the mouse tyrosine hydroxylase promoter (mTH) to drive Cre expression in VTA DA neurons. Intra-VTA co-infusion of AAV8-mTH-Cre with an AAV vector harboring a Cre-dependent yellow fluorescent protein expression cassette yielded high efficiency (82%) and high fidelity (73%) targeting of tyrosine hydroxylase-positive VTA neurons in C57BL/6J mice. Co-infusion of AAV8-mTH-Cre with a vector harboring a Cre-dependent channelrhodopsin (ChR2) expression cassette permitted optical regulation of VTA neurons with electrophysiological features consistent with VTA DA neurons. Moreover, C57BL/6J mice expressing ChR2 in VTA DA neurons rapidly acquired optical self-stimulation behavior. Thus, this AAV-only approach should facilitate investigation of VTA DA neuron contributions to reward-related behaviors and permit comparative assessments in reward circuit function in inbred and mutant mouse strains.

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• Synaptic dysfunction and adaptation after NMDA receptor ablation in the mouse medial prefrontal cortex

  Neuropsychopharmacology · 2026-03-14

  articleOpen access

  N-methyl-D-aspartate receptors (NMDARs) in the prefrontal cortex (PFC) are critical regulators of neuronal excitability, synaptic plasticity, and cognitive function. NMDAR disruptions, including pharmacological blockade and anti-NMDAR encephalitis, can mimic symptoms of schizophrenia. These observations support the glutamate hypothesis of schizophrenia, which posits that symptoms arise from abnormal corticolimbic glutamatergic signaling. Further evidence for this theory includes abnormal expression of NMDARs and decreased dendritic spine density in the PFC of individuals with schizophrenia, as well as altered spine density and synaptic transmission caused by genetic manipulation of NMDARs. However, it is unknown how progressive loss of NMDAR function in the PFC during adolescence-a developmental time period associated with symptom onset in schizophrenia -affects excitatory synaptic structure and function. In this study, we used in vivo genome editing to ablate expression of the Grin1 gene, which encodes the obligate GluN1 subunit of NMDARs, in medial PFC neurons of female and male adolescent mice. We assessed synaptic density and function in layer V pyramidal neurons using whole-cell patch-clamp electrophysiology, integrated with confocal imaging of dendritic spine architecture in recorded neurons. NMDAR ablation caused an early decrease in basilar dendritic spine density, followed by a rebound in spine density and a corresponding increase in AMPAR-mediated synaptic transmission. These effects of pan-neuronal NMDAR ablation were not observed after a more specific manipulation of excitatory neurons. Our findings demonstrate that NMDAR ablation triggers a cascading reorganization of local PFC networks, which may include compensatory processes that maintain allostasis but are impaired in disease states.

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• Global but not myeloid lineage-directed Girk3 deletion increases bone mass in female mice

  JBMR Plus · 2025-08-12

  articleOpen access

  Abstract Germline and osteoblast-directed deletion of G protein-gated inwardly rectifying K+ channel 3 (Girk3) was recently shown to increase bone mass after 18 wk of age in male mice. Here, we show that germline Girk3 deletion also increases trabecular and cortical bone mass and increases the mechanical strength of the femur in female mice after 18 wk of age. Unlike male mice, however, osteoblast-directed Girk3 deletion using 2.3 kb-Col1a1-Cre does not increase bone mass in adult female mice. To discover mechanisms underlying high bone mass in female Girk3−/− mice, bulk RNA-sequencing was performed on 2-d-old calvarial bone, revealing lower expression of proinflammatory cytokines such as IL-1β and IL-6 in Girk3−/− mice. Accordingly, cytokines and chemokines are largely suppressed in the circulation of adult Girk3−/− mice compared to WT littermates. The cytokines GM-CSF, IL-1β, IL-2, and IL-9 are reduced in the serum of both male and female Girk3−/− mice, while eotaxin, IFNγ, MIP-1α, and others are sexually dimorphic. Histomorphometry reveals that osteoclast activity is modestly reduced in Girk3−/− bone, which is supported by in vitro osteoclast resorption assays. However, deletion of Girk3 in myeloid-lineage cells with LysM-Cre is not sufficient to recapitulate high bone mass in either male or female mice. Moreover, female Girk3−/− mice are not protected from ovariectomy-induced bone loss. Finally, single-cell screening using cytometry by time-of-flight in the BM revealed no differences in immune cell abundances due to global Girk3 deletion. Taken together, while Girk3 regulates inflammatory cytokine expression in the bone and serum, deletion of Girk3 in myeloid-lineage cells does not affect bone mass.

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• Editorial: Advances in pharmacotherapy for alcohol use disorder: from mechanisms to clinical interventions

  Frontiers in Pharmacology · 2025-08-15

  editorialOpen access

  Pathophysiological adaptation in the striatum has been recognized as a principal brain dysfunction resulting from chronic alcohol exposure, specifically contributing to maladaptive reward-seeking behaviors (Corbit and Janak, 2016). Given the different roles of striatal subregions in shaping reward-seeking patterns and motivation, the detailed characterization of striatal adaptations to chronic alcohol consumption is important to understand the progression from voluntary to inflexible alcohol-seeking behaviors. Recent research by Duffus et al. employed mass spectrometry analysis of protein abundance in the dorsomedial, dorsolateral, and nucleus accumbens subregions of the striatum, at two different abstinence time points following chronic, voluntary alcohol drinking in male and female mice (Duffus et al., 2024). Their comprehensive proteomics data illuminated brain adaptations, such as changes in neurodegeneration-associated proteins, that differed as a function of abstinence duration, subregion, and sex. A general feature, however, was that chronic alcohol drinking appeared to primarily alter proteins important for neuronal structure and cellular health, rather than induce neuroinflammation in the striatum; proteomic profiling revealed changes in proteins and pathways associated with metabolic, cellular organization, protein translation, and molecular transport processes. These findings are in contrast to the literature linking alcohol dependence to upregulation of neuroinflammatory processes, suggesting that the role of neuroinflammation in alcohol-related behavioral changes may vary across different brain regions. On the other hand, it is not clear whether the chronic drinking model employed induces alcohol dependence, and discrepancies with prior work could also be related to differences in the degree of alcohol exposure or the abstinence time point at which proteins were measured. Nonetheless, this 40 work not only demonstrates the of subregion-specific characterization but also provides a 41 new list of proteins that establishes a foundation for future investigations of alcohol-induced changes 42 to striatal neuronal structure and cellular health their behavioral consequences. 43 Semaglutide, a long-acting analogue glucagon-like peptide-1, has emerged as an effective weight-44 loss treatment through appetite regulation in both diabetic and non-diabetic individuals. 45Accumulative evidence indicates its efficacy in weight loss-independent actions as well (Drucker, 46 2024). GLP-1 receptor agonists also have recently shown their potential as treatments for AUD in 47 rodent and nonhuman primate studies, and clinical trials to evaluate the efficacy of the agonists have 48 been initiated for AUD patients (Marty et al., 2020;Farokhnia et al., 2025;Hendershot et al., 2025). 49Aranas et al. examined, using rodent models in both male and female, the potential synergistic 50 effects of combining semaglutide with the well-known anti-smoking agents, varenicline or 51 bupropion, in the reduction of alcohol intake, while simultaneously assessing the impact of a high-fat 52 diet (HFD) (Aranas et al., 2023). Aranas et al. confirmed that semaglutide as a monotherapy 53 effectively reduced alcohol intake and preference. Notably, when semaglutide was combined with 54 either of varenicline, bupropion or HFD, it did not change the effects of semaglutide on the reduction 55 of alcohol intake, suggesting that pharmacological interventions to target GLP-1 provide sufficient 56 effects for AUD without requiring "complex combination regimens", offering the potential for 57 optimizing treatments for both AUD and obesity. Interestingly, HFD feeding in this study (Aranas et 58 al., 2023) was also found to be equally effective in reducing alcohol drinking, which is consistent 59 with several rodent studies. Considering the nutritional deficiencies following prolonged chronic 60 alcohol consumption and increased intake of palatable food in recovering patients, these data may 61 have important clinical implications in the management of AUD. While a complex relationship 62 between nutrition and AUD exists, the possibility of utilizing a non-pharmacological nutritional 63 intervention that could facilitate the pathway of recovery needs further investigation (White & Sirohi, 64 2024). 65Treatment availability for AUD continues to be limited, evident as less than 10% of patients obtain 66 any form of treatment, and even the treatment rates vary around the world (Venegas et al., 2021). 67This gap in treatment has prompted exploration of alternative medications, particularly in regions 68 where traditional practices are culturally relevant and accepted. For example, AUD and its 69 comorbidities pose significant public health challenges in Africa, where traditional medicines are also 70 suggested as a complementary and alternative option due to the limited availability of conventional 71 medications.

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• Cardiac Fibroblasts: Helping or Hurting

  Genes · 2025-03-27 · 8 citations

  reviewOpen access

  Cardiac fibroblasts (CFs) are the essential cell type for heart morphogenesis and homeostasis. In addition to maintaining the structural integrity of the heart tissue, muscle fibroblasts are involved in complex signaling cascades that regulate cardiomyocyte proliferation, migration, and maturation. While CFs serve as the primary source of extracellular matrix proteins (ECM), tissue repair, and paracrine signaling, they are also responsible for adverse pathological changes associated with cardiovascular disease. Following activation, fibroblasts produce excessive ECM components that ultimately lead to fibrosis and cardiac dysfunction. Decades of research have led to a much deeper understanding of the role of CFs in cardiogenesis. Recent studies using the single-cell genomic approach have focused on advancing the role of CFs in cellular interactions, and the mechanistic implications involved during cardiovascular development and disease. Arguably, the unique role of fibroblasts in development, tissue repair, and disease progression categorizes them into the friend or foe category. This brief review summarizes the current understanding of cardiac fibroblast biology and discusses the key findings in the context of development and pathophysiological conditions.

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• Designing allosteric modulators to change GPCR G protein subtype selectivity

  Nature · 2025-10-22 · 9 citations

  articleOpen access

  G-protein-coupled receptors (GPCRs) convert extracellular signals into intracellular responses by signalling through 16 subtypes of Gα proteins and two β-arrestin proteins. Biased compounds—molecules that preferentially activate a subset of these proteins—engage therapy-relevant pathways more selectively1 and promise to be safer, more effective medications than compounds that uniformly activate all pathways2. However, the determinants of bias are poorly understood, and we lack rationally designed molecules that select for specific G proteins. Here, using the prototypical class A GPCR neurotensin receptor 1 (NTSR1), we show that small molecules that bind to the intracellular GPCR–transducer interface change G protein coupling by subtype-specific and predictable mechanisms, enabling structure-guided drug design. We find that the intracellular, core-binding compound SBI-553 switches the G protein preference of NTSR1 through direct intermolecular interactions3–5, promoting or preventing association with specific G protein subtypes. Modifications to the SBI-553 scaffold produce allosteric modulators with distinct G protein selectivity profiles. Selectivity profiles are probe independent, conserved across species and translate to differences in activity in vivo. Our studies show that G protein selectivity can be tailored with small changes to a single chemical scaffold targeting the receptor–transducer interface. Moreover, given that this pocket is broadly conserved, our findings could provide a strategy for pathway-selective drug discovery that is applicable to the diverse GPCR superfamily. Studies of the G-protein-coupled receptor NTSR1 show that the G protein selectivity of this receptor can be modified by small molecules, enabling the design of drugs that work by switching receptor subtype preference.

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• Membrane-associated estrogen receptor α prevents the amyloid β-induced suppression of GIRK channel activity in hippocampal neurons from female mice

  Biology of Sex Differences · 2025-11-06

  articleOpen access

  Abstract Background Amyloid β oligomers (oAβ) are a key pathogenic driver in Alzheimer’s Disease (AD). Neuronal G protein-gated inwardly rectifying K + (GIRK/Kir3) channels are important regulators of neuronal excitability and prominent somatodendritic effectors for inhibitory G protein-coupled receptors, including the γ-aminobutyric acid type B receptor (GABA B R). We previously reported a male-specific suppression of GIRK channel activity in hippocampal (HPC) neurons evoked by oAβ in in vitro, ex vivo, and in vivo mouse models of AD, and showed that this adaptation correlated with synaptic and cognitive impairment. Using pharmacological approaches, we showed that this adaptation is mediated by co-activation of cellular prion protein (PrP C ) and metabotropic glutamate receptor 5 (mGluR5) and requires activation of cytosolic phospholipase A2 α (cPLA 2 α). However, the mechanisms underlying the sex specificity was unknown. Given the clinical context that females exhibit a 2-fold higher incidence of AD than males, and the loss of neuroprotective estrogen by menopause contributes to the sex differences in AD, we postulated that estrogen-associated resilience underlies this sex dimorphism of oAβ action. Methods To examine the strength of GIRK-dependent signaling in HPC neurons, we performed electrophysiology in primary HPC cultures from neonatal male and female mice and then measured whole-cell currents evoked by the direct-acting GIRK channel agonist ML297 and the GABA B R-selective agonist baclofen. We used an array of genetic and pharmacological approaches to investigate the molecular mechanism(s) underlying the vulnerability and resilience of GIRK channel activity to oAβ in male and female HPC neurons, respectively. Results We found that resilience to the oAβ-induced and PrP C /mGluR5-dependent suppression of GIRK channel activity in female HPC neurons is conferred by membrane-associated estrogen receptor α (mERα) and caveolin 1 (Cav1). When this resilience factor is blocked or absent, oAβ suppresses GIRK channel activity in female HPC neurons via the same PrP C -mGluR5-cPLA 2 α signaling pathway identified previously in male neurons. Conclusion As estrogen levels decline with aging and menopause, the protective influence of mERα/Cav1 may diminish, unmasking the oAβ-induced suppression of GIRK channel activity and exacerbating disease progression in females. Plain english summary While amyloid β plaques (Aβ) are notable hallmarks of Alzheimer’s Disease (AD), cognitive impairment in the early stages of the disease tracks more closely with the level of soluble Aβ oligomers (oAβ) in the brain. oAβ promotes cognitive deficits by disrupting the balance of excitatory and inhibitory influences on neurons in brain regions important for learning and memory such as the hippocampus, but the underlying molecular targets of oAβ and its pathogenic mechanisms are not fully understood. We recently demonstrated that oAβ weakens the activity of a prominent inhibitory influence on neuronal excitability (the GIRK channel) in the hippocampus of male but not female mice. This sexually dimorphic effect of oAβ was interesting and unexpected given that women are twice as likely to develop AD than men, and because disease progression is more aggressive in women. In this study, we investigated the mechanisms underlying the resilience of GIRK channels in female hippocampal neurons to oAβ. We found that resilience is conferred by estrogen and one of its receptors. When the influence of this receptor is diminished using pharmacological or genetic interventions, oAβ weakens GIRK channel activity in female and male neurons to a similar degree, and via the same mechanism. We speculate that with the onset of menopause, the protective influence of estrogen on GIRK channel activity in the hippocampus begins to wane. This, combined with other female-specific effects of oAβ on neuronal activity, contributes to the increased incidence and severity of AD in females.

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• Synaptic Dysfunction and Compensation After NMDA Receptor Ablation in the Mouse Medial Prefrontal Cortex

  bioRxiv (Cold Spring Harbor Laboratory) · 2025-08-17

  preprintOpen access

  N-methyl-D-aspartate receptors (NMDARs) in the prefrontal cortex (PFC) are critical regulators of neuronal excitability, synaptic plasticity, and cognitive function. NMDAR disruptions, including pharmacological blockade and anti-NMDAR encephalitis, can mimic symptoms of schizophrenia. These observations support the glutamate hypothesis of schizophrenia, which posits that symptoms arise from abnormal corticolimbic glutamatergic signaling. Further evidence for this theory includes abnormal expression of NMDARs and decreased dendritic spine density in the PFC of individuals with schizophrenia, as well as altered spine density and synaptic transmission caused by genetic manipulation of NMDARs. However, it is unknown how progressive loss of NMDAR function in the PFC during adolescence - a developmental time period associated with significant synaptic pruning and symptom onset in schizophrenia - affects excitatory synaptic structure and function. In this study, we used in vivo genome editing to ablate expression of the Grin1 gene, which encodes the obligate GluN1 subunit of NMDARs, in medial PFC neurons of female and male adolescent mice. We assessed synaptic density and function in layer V pyramidal neurons at multiple time points using whole-cell patch-clamp electrophysiology, integrated with confocal imaging of dendritic spine architecture in recorded neurons. NMDAR ablation caused an early decrease in basilar dendritic spine density, followed by a rebound in spine density and corresponding increase in AMPAR-mediated synaptic transmission, suggesting that synaptic compensation maintains an allostatic set point. Our findings demonstrate that NMDAR ablation initially disrupts local PFC networks, followed by recovery via compensatory processes that could be impaired in disease states.

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• Chronic ethanol exposure in mice evokes pre‐ and postsynaptic deficits in GABAergic transmission in ventral tegmental area GABA neurons

  British Journal of Pharmacology · 2024-10-02 · 6 citations

  articleOpen access

  Abstract Background and purpose GABAergic neurons in mouse ventral tegmental area (VTA) exhibit elevated activity during withdrawal following chronic ethanol exposure. While increased glutamatergic input and decreased GABA A receptor sensitivity have been implicated, the impact of inhibitory signaling in VTA GABA neurons has not been fully addressed. Experimental approach We used electrophysiological and ultrastructural approaches to assess the impact of chronic intermittent ethanol vapour exposure in mice on GABAergic transmission in VTA GABA neurons during withdrawal. We used CRISPR/Cas9 ablation to mimic a somatodendritic adaptation involving the GABA B receptor (GABA B R) in ethanol‐naïve mice to investigate its impact on anxiety‐related behaviour. Key results The frequency of spontaneous inhibitory postsynaptic currents was reduced in VTA GABA neurons following chronic ethanol treatment and this was reversed by GABA B R inhibition, suggesting chronic ethanol strengthens the GABA B R‐dependent suppression of GABAergic input to VTA GABA neurons. Similarly, paired‐pulse depression of GABA A receptor‐dependent responses evoked by optogenetic stimulation of nucleus accumbens inputs from ethanol‐treated mice was reversed by GABA B R inhibition. Somatodendritic currents evoked in VTA GABA neurons by GABA B R activation were reduced following ethanol exposure, attributable to the suppression of GIRK (K ir 3) channel activity. Mimicking this adaptation enhanced anxiety‐related behaviour in ethanol‐naïve mice. Conclusions and implications Chronic ethanol weakens the GABAergic regulation of VTA GABA neurons in mice via pre‐ and postsynaptic mechanisms, likely contributing to their elevated activity during withdrawal and expression of anxiety‐related behaviour. As anxiety can promote relapse during abstinence, interventions targeting VTA GABA neuron excitability could represent new therapeutic strategies for treatment of alcohol use disorder.

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Frequent coauthors

• Kevin Wickman

  University of Minnesota

  51 shared

• Allison Anderson

  University of Minnesota

  22 shared

• Baovi N. Vo

  University of Minnesota

  22 shared

• Corey R. Hopkins

  Nebraska Medical Center

  19 shared

• C. David Weaver

  Vanderbilt University

  19 shared

• Raquel E. Rodrı́guez

  15 shared

• Gemma Arsequell

  Institute of Advanced Chemistry of Catalonia

  8 shared

• Iván Rodríguez-Martín

  Universidad Francisco de Vitoria

  7 shared