Developing effective therapeutics for pain requires selecting endpoints that accurately reflect the biological, functional, and neurophysiological changes in a model. High-quality study design relies on capturing data across multiple facets of disease expression, as no single endpoint can fully represent the complexity of pain. Across preclinical pain research, a combination of complementary endpoints, such as behavioral assessments, biomarker evaluations, and electrophysiological measurements, helps support strong, translational study design.
Behavioral readouts remain central to evaluating hypersensitivity, spontaneous pain, and functional impairment. Traditional measures, such as von Frey filaments, mechanical/tactile allodynia scoring, or thermal nociceptive tests, provide quantifiable readouts of sensory processing and allow researchers to detect shifts in pain thresholds with high sensitivity.
However, pain manifests far beyond stimulus-evoked responses. Additional functional assessments such as open-field exploration, gait analysis, activity monitoring, and weight-bearing distribution provide a broader understanding of an animal model's overall condition. In both rodent and large-animal models (such as pigs), altered stride patterns, reduced exploratory behavior, movement avoidance, or asymmetric stance often correlate closely with underlying nociceptive states.
These measures allow therapeutic candidates to be evaluated on their ability to improve integrated functional behaviors, an increasingly important aspect of translational relevance as researchers look for evidence that preclinical improvements may extend to human outcomes.
Biomarkers provide objective, quantifiable indicators of biological processes occurring within a model and offer essential mechanistic insight. As measurable signatures in tissue, blood, CSF, or other biofluids, biomarkers help identify inflammatory cascades, neurodegenerative mechanisms, metabolic stress, tissue integrity, and therapeutic modulation.
A range of analytical and imaging tools support detailed biomarker evaluation:
Flow cytometry: Characterizes immune cell populations, activation states, and changes in inflammatory environments.
Multiplex cytokine/chemokine analysis: Measures multiple inflammatory mediators simultaneously, allowing rapid detection of shifts in biological pathways.
Immunohistochemistry (IHC): Identifies protein expression and localization, including markers of neuroinflammation, axonal injury, glial activation, and regenerative activity.
Histological analysis: Evaluates tissue-level changes such as nerve fiber density, edema, fibrosis, Wallerian degeneration, or structural remodeling.
Blood flow imaging: Assesses vascular involvement, perfusion deficits, or ischemia-associated tissue responses.
Together, these assays help characterize the biological pathways active within a model and support the interpretation of therapeutic mechanisms of action. MD Biosciences works with sponsors to select relevant biomarkers and assays tailored to their compound, disease area, and study goals.
Electrophysiological readouts provide direct, quantitative insight into neural pathway function and are especially valuable in models of neuropathic pain, neurodegeneration, or peripheral nerve injury. Because they detect changes in conduction and signaling before behavioral recovery occurs, electrophysiological endpoints offer early, objective evidence of therapeutic impact. Common techniques include:
Sensory and motor evoked potentials (SEPs/MEPs): Assess cortical and spinal responsiveness to controlled stimuli.
Nerve conduction velocity (NCV): Evaluates the speed and efficiency of peripheral nerve signaling.
Compound action potentials (CAPs): Measure the summed electrical activity of nerve fibers.
Electromyography (EMG): Provides insight into neuromuscular signaling integrity and muscle activation patterns.
Electrophysiology bridges the gap between functional outcomes and underlying neural physiology, supporting mechanistic interpretation and enhancing the translational strength of study findings. View our whitepaper here for more information on electrophysiology.
Integrating Endpoints for Greater Translational Strength
Pain is multifactorial, and no single endpoint captures its full complexity. Relying solely on behavioral or stimulus-evoked measures can overlook important biological or functional changes occurring within a model. By combining behavioral assessments with molecular biomarkers, neural imaging, histology, and electrophysiological readouts, studies generate a more layered and reliable dataset. This improves reproducibility, reduces the likelihood of false-positive interpretation, strengthens mechanistic understanding, and provides greater confidence in therapeutic effects as compounds advance through development. This integrative approach is particularly valuable in neurological diseases, where motor, sensory, cognitive, and inflammatory elements often intersect. Bringing together functional, physiological, and biochemical measures offers a more complete understanding of disease processes and therapeutic potential.
Contact one of our scientists today to discuss pain studies in more detail.