Pain & Epilepsy

InterVivo offers a capability for assessment of NCEs in rodent tests of pain and epilepsy. Several drug classes prevent both experimentally-induced seizures and reduce pain in animal models which translate into the clinic. Measurement of drug effects across both model types enables a broadening of therapeutic indications, and increased knowledge about biological property.

Chronic pain is typically sub-divided into pain of inflammatory and neuropathic origin. Due to distinct etiologies, both types of pain respond differently to certain drug classes and specific models have been developed for each type.

The formalin test has both an inflammatory and neuropathic pain component and thus serves as a useful initial screening test. Intra-plantar injection of formalin produces a biphasic incidence of licking the injected area, with an early (i.e. 0-10min) phase due to direct activation of TRPA1 channels localized on sensory neurons, and a late (10-60min) phase which is believed to reflect both central sensitization and local inflammatory response to formalin-induced TRPA1 channel activation and local tissue damage.

The withdrawal latency to a radiant heat source provides a very simple measure to an acute stimulus. This test is used to evaluate treatments for the treatment of acute pain.

Intradermal injection of the mast cell degranulator Compound 48/80 produces a local release of histamine and intense scratching of the injected area. Since histamine elicits itch in humans this response is believed to reflect a similar sensation in mice.

The intra-plantar injection of the polysaccharide carrageenan produces an acute inflammatory response that peaks approximately 3-4h post injection. Edema is measured by paw volume, and pain is measured by withdrawal response to paw pressure (Randall-Selitto). Drugs such as the NSAID indomethacin, and COX2 inhibitors such as celecoxib reliably limit the edema and hyperalgesia produced by carrageenan.

Validation Data: Comparison of NSAID (Indomethacin) and COX2 inhibitor (Celecoxib) in the Carrageenan model.

Male rats were injected with Carrageenan into the plantar surface of hindpaw (RHS). 4h later, (A) paw volume (plethysmometer), (B) paw pressure (Randall-Selitto) were measured. At the study completion,(C) GI and stomachs were assessed histologically. Both indomethacin (10mg/kg, oral) and celecoxib (10mg/kg, oral) reduced paw volume and hyperalgesia score relative to vehicle controls.

Indomethacin, but not celecoxib, produced clear signs of GI lesions, with epithelial erosion and microhaemorrhage evident.

Transection of the tibial and peroneal branches of the sciatic nerve produces a long lasting neuropathy of the denervated paw, characterized by tactile and cold allodynia. These effects may be maintained for several weeks post surgery enabling long term chronic studies, and/or within subject test designs to be undertaken.

Validation Data: Effect of Tegretol® (Carbamazepine) against allodynia and hyperalgesia following SNI

SNI and sham operated rats were surgically prepared. By 10 days all SNI lesioned animals developed tactile/cold allodynia relative to sham controls. Approximately 30 days post surgery the animals were treated with Carbamazepine (CBZ) 10-30mg/kg. Acute CBZ attenuated the thermal hyperalgesia and cold allodynia, but not the tactile allodynia

The intra-plantar injection of Freund's Complete Adjuvant (FCA) produces an acute inflammatory response that peaks at approximately 24h, and is maintained for over 30 days post injection. Edema is measured by by the threshold withdrawal response to Von Frey hairs. NCEs can be tested either acutely (i.e. 1 day post FCA), or longer term (i.e. 30 days post FCA). An advantage of testing longer term FCA injection is that this may more closely mimic the deleterious effects of long term inflammatory conditions such as rheumatoid arthritis.

Validation Data: Timecourse of paw oedema,tactile allodynia and histology following intra-planter injection of Freunds adjuvant.

Rats were injected intra-plantar with Freunds adjuvant (0.5-1mg/ml, 50uL volume) and paw volume measured over 30 days. At 30 day (but not 1 day), some evidence of bone erosion (*) and cartilage loss (#) was evident in some animals.

A 1cm incision through the plantar surface of the hindpaw produces a sensitization to a tactile stimulus applied 1 to 2 days post surgery. NCE’s may be tested against the withdrawal response to this tactile response either 1 or 2 days post surgery. This model is used to evaluate treatments for post operative pain.

Acute seizure tests include maximal electroshock (MES), the chemical convulsant pentylenetetrazol (s.c.PTZ), and the 6Hz model of psychomotor seizure. Similar to pain, epilepsy is a heterogeneous condition which responds differently to drugs. Assessment of an NCE across the MES, PTZ and 6Hz models provides a drug signature and insight as to which seizure type may be treatable. Together these 3 acute tests identify all known AED's, and the MES and PTZ tests are currently recognized as the gold standard early phase screening tests1.

Validation Data: Comparison of phenytoin, valproate and levetiracetam against MES, s.c.PTZ and 6Hz-induced seizures in the mouse.
Dose response curves for each drug against each seizure test. Note each drug has a unique profile across these tests, consistent with distinct mechanism of action and clinical profile.

Repeated subconvulsive electrical stimulation of discrete CNS structures (typically amygdala, hippocampus) will lead to induction (kindling) of overt seizures. As such this may model human partial seizures with secondary generalization. Typical endpoints are (a) behavioral assessment of seizure intensity (Racine scale), and (b) electrophysiological assessment of afterdischarge EEG. NCEs may be tested either as a pretreatment to examine a block of an already established kindled seizure, or administered during kindling development to assess antiepileptogenic potential. Today the kindling model is the only chronic model widely used in most AED screening programs1.

Validation Data: Effect of levetiracetam (Keppra ®) against amygdala kindled seizures
Amygdala seizures

In rats fully kindled to stage 5 seizures and with afterdischarge duration typically in the range of 100-120s, the acute pretreatment of levetiracetam (30-60mg/kg) dose dependently reduced EEG afterdischarge duration, and seizure intensity. Clinically, Keppra® is one of the most widely prescribed treatments for partial seizures.

1 Bialer & White (2010) Key factors in the discovery and development of new antiepileptic drugs. Nature Rev. Drug Discovery 9: 68-82.