Alzheimer's disease (AD) is a neurodegenerative disorder characterized by progressive memory loss with deterioration of cognitive functions. Recent data from the literature show that approximately 45.8% of people living with AD suffer from neuropathic pain and this untreated pain continues to be a serious public health problem. However, unfortunately, our understanding of its processing is still very incomplete due to its high complexity, especially for special populations, which should deserve more attention and respect, such as people affected by AD, and who in the vast majority of cases cases are unable to verbalize that they feel pain. Furthermore, the literature is not clear about which type of neuropathic pain these patients are affected by. Therefore, our hypothesis is that it is central neuropathic pain (CP), defined as a lesion in the central nervous system leading the individual to develop hypersensitivity conditions such as allodynia. Hemopressin (Hp) family peptides are a tool with great potential for the treatment of neuropathic pain and AD, given their ability to modulate the activity of the endocannabinoid system via specific receptors, such as the type 1 cannabinoid receptor (CB1R). It has already been demonstrated that Hp (CB1R inverse agonist) and VD-Hp (CB1R agonist) have an antinociceptive effect in other models, but little is known about this action in CP associated with AD. Experimental models are fundamental to characterize, in the biological complexity of laboratory animals, diseases of the most varied origins that, in addition to causing a negative impact, drastically reduce the quality and life expectancy of the population. In the absence of laboratory studies that characterize these two disorders already mentioned above, the objective of this study is to standardize an animal model that makes it possible to understand not only the behavioral and physiological complexity of the relationship between CP and AD, but also to propose a way to reverse this situation by generating antinociception.

 

In order to mimic the characteristic neurodegeneration of sporadic AD, intracerebroventricular (icv) injection of streptozotocin (STZ) was used, a method well established in the literature where neuronal death occurs due to an interruption of cellular glucose metabolism, causing progressive deficit in memory, learning and long-term cognitive behavior. Therefore, male Wistar rats (90 days, 250-350 kg) were submitted to bilateral icv injections (4µL). To standardize the model, the animals' mechanical, thermal and cold sensitivity were evaluated using von Frey monofilaments, acetone spray test and hot plate on the 3rd, 7th, 10th and 14th days after surgery, as well as the cognitive assessment through the object recognition test on the 5th and 6th days together with the open field test (initial 3 minutes of the first habituation phase) investigating whether the animals had any anxiety/depression-like behavior. To evaluate the antinociceptive effect of VD-Hp and Hp, they were administered orally (gavage) on the 7th day after surgery, with mechanical sensitivity being evaluated 1, 3 and 6 hours after administration.

 

The results referring to the standardization of the model showed that the STZ animals developed cognitive impairment in short-term memory (p=0.0003; total exploration time: p=0.0373) and long-term memory (p=0.0002; total exploration time: p=0.0315). Assessment of anxiety/depression-like behavior showed no significant difference between groups in general activity (p=0.2947) and amount of fecal bolus produced (p=0.4205). However, a preference was observed for both the CT (p=0.0037) and STZ (p=0.0127) groups for the periphery, in addition to the STZ group showing a shorter time on two paws (p=0.0411). The administration of STZ icv induced the development of mechanical sensitivity (CT n=13; STZ n=13) [3, 7, 10 and 14 (p<0.0001) days], thermal to cold (CT n=9; STZ n=9) [3, 7, 10 and 14 (p<0.0001) days] and hot (CT n=4; STZ n=5) [3, 7, 10 and 14 (p<0.0001) days] observed throughout the evaluation period. Finally, we demonstrated that VD-Hp (3h [p =0.0105); 6h [p=0.0088]) and Hp (1h [p=0.0170]; 3h [p=0.0157]) reversed the mechanical sensitivity of the animals when evaluated 1, 3, 6h after its administration.

 

In conclusion, the presented results standardize the CP model associated with neurodegeneration in rats after STZ administration, accompanied by the development of cognitive deficit related to short and long term memory, the presence of some anxiety/depression-like behavior, mechanical and thermal hypersensitivity to cold and hot. In addition, the antinociceptive effect of VD-Hp and Hp was demonstrated, reversing the mechanical hypersensitivity of the animals.

 

ANDERSON, Alison R. et al. Policy implications for pain in advanced Alzheimer's disease. Pain Management Nursing, v. 22, n. 1, p. 3-7, 2021.

 

VAN KOOTEN, Janine et al. A review of pain prevalence in alzheimer's, vascular, frontotemporal and lewy body dementias. Dementia and geriatric cognitive disorders, v. 41, n. 3-4, p. 220-232, 2016.

 

MA, Chao. Animal models of pain. International anesthesiology clinics, v. 45, n. 2, p. 121-132, 2007.

 

WATSON, James C.; SANDRONI, Paola. Central neuropathic pain syndromes. In: Mayo clinic proceedings. Elsevier, 2016. p. 372-385.

 

HEIMANN, Andrea S. et al. Hemopressin as a breakthrough for the cannabinoid field. Neuropharmacology, v. 183, p. 108406, 2021.

 

GRIEB, Paweł. Intracerebroventricular streptozotocin injections as a model of Alzheimer’s disease: in search of a relevant mechanism. Molecular neurobiology, v. 53, p. 1741-1752, 2016.

 

LANNERT, Heinrich; HOYER, Siegfried. Intracerebroventricular administration of streptozotocin causes long-term diminutions in learning and memory abilities and in cerebral energy metabolism in adult rats. Behavioral neuroscience, v. 112, n. 5, p. 1199, 1998.

 

CRUNFLI, Fernanda et al. Cannabinoid receptor type 1 agonist ACEA improves cognitive deficit on STZ-induced neurotoxicity through apoptosis pathway and NO modulation. Neurotoxicity Research, v. 35, p. 516-529, 2019.

 

The results obtained in this work are essential so that, through new therapeutic methods, a better quality of life is provided for patients affected by AD who suffer from neuropathic pain.

 

The animals used in this study were provided by the bioterium network of the Institute of Biomedical Sciences of the University of São Paulo (ICB-USP). The experiments were conducted in accordance with the rules of the Brazilian College of Animal Experimentation (COBEA) and the guidelines of the National Council for the Control of Animal Experimentation (CONCEA) with an experimental protocol approved by the Committee on Ethics in the Use of Animals (CEUA) of the Instituto de Biomedical Sciences at the University of São Paulo under protocol nº 2532260421.