Ex Vivo Study of Colon Health, Contractility and Innervation in Male and Female Rats after Regular Exposure to Instant Cascara Beverage
"> Figure 1
<p>Experimental procedure. Male and female rats were given either water (control group) or IC beverage for a duration of 4 weeks. During the fourth week, animals were sacrificed to analyze the health of the colon wall using histological methods, colonic muscle strip contractility employing organ bath procedures, and its innervation using immunohistochemistry techniques.</p> "> Figure 2
<p>Schematic representation of how the longitudinal and circular muscle strips were obtained. Once the colonic segment was removed from the rat, it was pinned on a Petri dish covered with Sylgard<sup>®</sup> and filled with Krebs solution. A longitudinal cut was performed through the mesenteric border. Once stretched and pinned on the dish surface, the mucosa and submucosa layers were removed, and the circular (CM) and longitudinal (LM) muscle strips were obtained by cutting perpendicular or parallel to the longitudinal axis of the colon, respectively.</p> "> Figure 3
<p>Experimental protocol of the organ bath experiments. Longitudinal and circular muscle strips were suspended in organ bath cups. Upper panel: After an initial 60 min stabilization period with 3 Krebs renewals, potassium chloride (KCl) was added at 50 mM to study the contractility of the strips. (<b>A</b>) After 2 Krebs renewals, the strips were electrically stimulated (EFS) at increasing frequencies (0.1–20 Hz) and posteriorly with acetylcholine (ACh) at increasing concentrations (10<sup>−8</sup>–10<sup>−5</sup> M), Krebs was renewed two times before the next concentration was added. Lower panel (<b>B</b>): The same electrical and chemical stimulations were repeated in the presence of atropine (10<sup>−6</sup> M), only this time just with ACh 10<sup>−5</sup> M and without Krebs renewals. Finally, Krebs was renewed twice, and KCl (50 mM) was added to the organ bath.</p> "> Figure 4
<p>Representative traces of colonic smooth muscle contractile responses and parameters measured. (<b>A</b>) Measurement of the amplitude of the phasic (PA) and tonic (TA) components of the contractions induced by ACh (and KCl). TA occurs after PA, as a plateau, generally below the value of PA. (<b>B</b>) Measurement of Amplitude 1 (A<sub>1</sub>) and Amplitude 2 (A<sub>2</sub>) of the contraction induced during and after electrical stimulation (EFS), respectively. Thick vertical arrow in A and thin double-head arrow in B represent stimulus (administration of ACh in (<b>A</b>), EFS duration in (<b>B</b>)).</p> "> Figure 5
<p>The impact of Instant Cascara (IC) beverage regarding the microscopic features of the colon in male and female rats. Colonic damage (<b>A</b>) was evaluated by examining ten randomly selected fields per section at 40× magnification, using three distinct sections of colon tissue for each animal. In addition, the width of the muscle layer was measured (<b>B</b>). The images (<b>C</b>–<b>F</b>) show the muscle layer of the colon. During the fourth week of IC beverage administration, tissue samples were collected from animals across four experimental groups: Males—Control, Males—IC, Females—Control, and Females—IC. Each group consisted of six animals. The results are presented as mean ± SEM (standard error of the mean). Significant differences related to sex were observed, with <span class="html-italic">p</span> < 0.0001 indicated by #### for comparisons between Females—Control and Males—Control, and <span>$</span><span>$</span><span>$</span><span>$</span> for comparisons between Females—IC and Males—IC. Statistical analysis was conducted using one-way ANOVA with Bonferroni’s post hoc test. Bar: 100 μm.</p> "> Figure 6
<p>Electrical field stimulation (EFS) was applied to longitudinal (LM) and circular (CM) muscle strips using 10 s pulse trains (0.3 ms) at frequencies of 0.1, 0.5, 1, 2, 5, 10, and 20 Hz. The following metrics were recorded: (<b>A</b>) maximum amplitude observed during stimulation (Amplitude 1) in LM; (<b>B</b>) maximum amplitude detected after stimulation (Amplitude 2) in LM; (<b>C</b>) Amplitude 1 in CM; (<b>D</b>) Amplitude 2 in CM. Data are reported as mean ± SEM, with each group consisting of 5–6 rats and 17–20 muscle strips for both LM and CM. Statistically significant differences related to sex are indicated by the following: # <span class="html-italic">p</span> < 0.05; ## <span class="html-italic">p</span> < 0.01; ### <span class="html-italic">p</span> < 0.001; #### <span class="html-italic">p</span> < 0.0001 for Male—Control vs. Female—Control comparisons; <span>$</span> <span class="html-italic">p</span> < 0.05; <span>$</span><span>$</span><span>$</span><span>$</span> <span class="html-italic">p</span> < 0.0001 for Male—IC vs. Female—IC comparisons. Statistical significance was determined using two-way ANOVA followed by Bonferroni’s post hoc test.</p> "> Figure 7
<p>The effects of electrical field stimulation (EFS) on longitudinal (LM) and circular (CM) muscle strips were evaluated under non-muscarinic conditions after atropine (10<sup>−6</sup> M). EFS comprised 10 s pulse sequences (0.3 ms, 100 V) across frequencies of 0.1, 0.5, 1, 2, 5, 10, and 20 Hz. The following were recorded: (<b>A</b>) peak amplitude during stimulation (Amplitude 1) in LM; (<b>B</b>) peak amplitude post stimulation (Amplitude 2) in LM; (<b>C</b>) Amplitude 1 in CM; (<b>D</b>) Amplitude 2 in CM. Data are reported as mean ± SEM for each experimental condition, with 5–6 rats and 17–20 muscle strips per condition. Sex-related significant differences are indicated as follows: # <span class="html-italic">p</span> < 0.05; ## <span class="html-italic">p</span> < 0.01; ### <span class="html-italic">p</span> < 0.001 comparing Male—Control to Female—Control; <span>$</span><span>$</span> <span class="html-italic">p</span> < 0.01; <span>$</span><span>$</span><span>$</span> <span class="html-italic">p</span> < 0.001 comparing Male—IC to Female—IC. Significant beverage-related differences are shown by + <span class="html-italic">p</span> < 0.05 for Females—IC versus Females—Control. Analysis was performed using two-way ANOVA with Bonferroni’s post hoc test.</p> "> Figure 8
<p>The contractile responses of longitudinal (LM) and circular (CM) muscle strips to acetylcholine (ACh) stimulation were examined by exposing the samples to a range of ACh concentrations (10<sup>−8</sup> to 10<sup>−5</sup> M). The data illustrate the phasic (<b>A</b>) and tonic (<b>B</b>) responses in LM and the phasic (<b>C</b>) and tonic (<b>D</b>) responses in CM. Results are presented as mean ± SEM for each group, with six rats per group and 17–20 strips per group for both LM and CM. The analysis was conducted using two-way analysis of variance (ANOVA) with subsequent Bonferroni’s correction for multiple comparisons, and no significant differences were observed (<span class="html-italic">p</span> > 0.05).</p> "> Figure 9
<p>Contractile responses of longitudinal (LM) and circular (CM) muscle strips to acetylcholine (ACh, 10<sup>−5</sup> M) were evaluated both before (−) and after (+) atropine (10<sup>−6</sup> M) administration. This setup was used to determine the role of muscarinic receptors in the ACh-induced contractions. The amplitude of the contractile responses, including both the phasic (PA) and tonic (TA) components, were compared for LM and CM under these conditions, specifically: (<b>A</b>) phasic response (PA) in LM; (<b>B</b>) tonic response (TA) in LM; (<b>C</b>) phasic response (PA) in CM; (<b>D</b>) tonic response (TA) in CM. Data were expressed as mean ± SEM for each group, with six rats and 17–20 strips per group for both LM and CM. Significant differences between atropine-treated and non-treated groups are indicated by ᵒᵒᵒᵒ <span class="html-italic">p</span> < 0.0001. Statistical evaluation was performed using two-way analysis of variance (ANOVA) followed by Bonferroni’s post hoc test.</p> "> Figure 10
<p>Impact of Instant Cascara (IC) beverage on the immunoreactivity of substance P (SP) within the colonic myenteric ganglia of male and female rats. Image (<b>A</b>) shows the myenteric ganglia of the colon, where SP is found. Images (<b>B</b>–<b>E</b>) show the myenteric ganglia immunoreactive to SP in the different experimental groups. The area immunoreactive to SP in the myenteric ganglia of the colon sections was quantified with the program Image J-Fiji (<b>F</b>). After a four-week period of administering the IC beverage, animals were sacrificed to evaluate various parameters across four experimental groups classified by sex and beverage type: Males—Control, Males—IC, Females—Control, and Females—IC. Results are expressed as mean ± SEM (standard error of the mean) for each group, with six animals per group. Statistical analysis was performed using a one-way analysis of variance (ANOVA), followed by Bonferroni’s post hoc test, with no significant differences found (<span class="html-italic">p</span> > 0.05). Bar: 50 μm.</p> "> Figure 11
<p>Whole-mount longitudinal muscle–myenteric plexus (LMMP) preparations were processed with the pan-neuronal marker HuC/D to detect all neurons (<b>A</b>) and neurons expressing neuronal nitric oxide synthase (nNOS), which is mainly present in myenteric neurons involved in inhibitory motor circuits (<b>B</b>). Using the program ImageJ-Fiji, the area occupied by the ganglia was measured (dashed lines (<b>A</b>)). Neurons immunoreactive for each of the mentioned markers were counted (<b>A</b>,<b>B</b>). Asterisk: extraganglionic neuron. White arrow: neuron positive for HuC/D and nNOS. Empty arrow: neuron positive for HuC/D and negative for nNOS. Scale bar: 100 µm.</p> ">
Abstract
:1. Introduction
2. Resources and Procedures
2.1. Ethical Declaration
2.2. Instant Cascara Beverage
2.3. Experimental Groups and Study Animals
2.4. Methodological Approach
2.5. Colon Wall Health: Histological Analysis
2.6. Colon Contractility: Organ Bath Study
2.7. Colonic Innervation: Immunohistochemistry Analysis
2.7.1. Colonic Sections
2.7.2. Whole-Mount Longitudinal Muscle–Myenteric Plexus Preparations
2.8. Drugs and Reagents
2.9. Data Evaluation
3. Results
3.1. General Health
3.2. Colon Wall Health: Histological Analysis
3.3. Colon Contractility: Organ Bath Study
3.3.1. Responses to KCl Stimulation
3.3.2. Electrical Field Stimulation
3.3.3. Chemical Stimulation with Acetylcholine
3.4. Colonic Innervation: Immunohistochemistry Analysis
3.4.1. Substance P
3.4.2. Myenteric Plexus Study in Whole-Mount LMMP Preparations
4. Discussion
4.1. Colon Wall Health
4.2. Colon Contractility
4.3. Colonic Innervation
4.4. Significance, Strengths and Limitations
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
- Sensoy, I. A Review on the Food Digestion in the Digestive Tract and the Used in Vitro Models. Curr. Res. Food Sci. 2021, 4, 308–319. [Google Scholar] [CrossRef] [PubMed]
- Suganya, K.; Koo, B.S. Gut–Brain Axis: Role of Gut Microbiota on Neurological Disorders and How Probiotics/Prebiotics Beneficially Modulate Microbial and Immune Pathways to Improve Brain Functions. Int. J. Mol. Sci. 2020, 21, 7551. [Google Scholar] [CrossRef] [PubMed]
- Furness, J.B. The Enteric Nervous System: Normal Functions and Enteric Neuropathies. Neurogastroenterol. Motil. 2008, 20, 32–38. [Google Scholar] [CrossRef] [PubMed]
- Spencer, N.J.; Hu, H. Enteric Nervous System: Sensory Transduction, Neural Circuits and Gastrointestinal Motility. Nat. Rev. Gastroenterol. Hepatol. 2020, 17, 338. [Google Scholar] [CrossRef] [PubMed]
- Nguyen, T.T.; Baumann, P.; Tüscher, O.; Schick, S.; Endres, K. The Aging Enteric Nervous System. Int. J. Mol. Sci. 2023, 24, 9471. [Google Scholar] [CrossRef] [PubMed]
- Holland, A.M.; Bon-Frauches, A.C.; Keszthelyi, D.; Melotte, V.; Boesmans, W. The Enteric Nervous System in Gastrointestinal Disease Etiology. Cell. Mol. Life Sci. 2021, 78, 4713. [Google Scholar] [CrossRef] [PubMed]
- Carabotti, M.; Scirocco, A.; Maselli, M.A.; Severi, C. The Gut-Brain Axis: Interactions between Enteric Microbiota, Central and Enteric Nervous Systems. Ann. Gastroenterol. Q. Publ. Hell. Soc. Gastroenterol. 2015, 28, 203. [Google Scholar]
- Liu, L.; Huh, J.R.; Shah, K. Microbiota and the Gut-Brain-Axis: Implications for New Therapeutic Design in the CNS. EBioMedicine 2022, 77, 103908. [Google Scholar] [CrossRef]
- Karakan, T.; Ozkul, C.; Akkol, E.K.; Bilici, S.; Sobarzo-Sánchez, E.; Capasso, R. Gut-Brain-Microbiota Axis: Antibiotics and Functional Gastrointestinal Disorders. Nutrients 2021, 13, 389. [Google Scholar] [CrossRef]
- Rao, M.; Gershon, M.D. The Bowel and beyond: The Enteric Nervous System in Neurological Disorders. Nat. Rev. Gastroenterol. Hepatol. 2016, 13, 517. [Google Scholar] [CrossRef]
- Mittal, R.; Debs, L.H.; Patel, A.P.; Nguyen, D.; Patel, K.; O’Connor, G.; Grati, M.; Mittal, J.; Yan, D.; Eshraghi, A.A.; et al. Neurotransmitters: The Critical Modulators Regulating Gut-Brain Axis. J. Cell Physiol. 2017, 232, 2359. [Google Scholar] [CrossRef] [PubMed]
- Margolis, K.G.; Cryan, J.F.; Mayer, E.A. The Microbiota-Gut-Brain Axis: From Motility to Mood. Gastroenterology 2021, 160, 1486. [Google Scholar] [CrossRef]
- Iriondo-DeHond, A.; Elizondo, A.S.; Iriondo-DeHond, M.; Ríos, M.B.; Mufari, R.; Mendiola, J.A.; Ibañez, E.; del Castillo, M.D. Assessment of Healthy and Harmful Maillard Reaction Products in a Novel Coffee Cascara Beverage: Melanoidins and Acrylamide. Foods 2020, 9, 620. [Google Scholar] [CrossRef] [PubMed]
- López-Parra, M.B.; Gómez-Domínguez, I.; Iriondo-DeHond, M.; Villamediana Merino, E.; Sánchez-Martín, V.; Mendiola, J.A.; Iriondo-DeHond, A.; del Castillo, M.D. The Impact of the Drying Process on the Antioxidant and Anti-Inflammatory Potential of Dried Ripe Coffee Cherry Pulp Soluble Powder. Foods 2024, 13, 1114. [Google Scholar] [CrossRef] [PubMed]
- Lachenmeier, W.; Sánchez-Martín, V.; López-Parra, M.B.; Iriondo-Dehond, A.; Haza, A.I.; Morales, P.; Dolores Del Castillo, M. Instant Cascara: A Potential Sustainable Promoter of Gastrointestinal Health. Proceedings 2023, 89, 21. [Google Scholar] [CrossRef]
- Iriondo-Dehond, A.; Uranga, J.A.; Del Castillo, M.D.; Abalo, R. Effects of Coffee and Its Components on the Gastrointestinal Tract and the Brain–Gut Axis. Nutrients 2020, 13, 88. [Google Scholar] [CrossRef] [PubMed]
- Romaní-Pérez, M.; Bullich-Vilarrubias, C.; López-Almela, I.; Liébana-García, R.; Olivares, M.; Sanz, Y. The Microbiota and the Gut–Brain Axis in Controlling Food Intake and Energy Homeostasis. Int. J. Mol. Sci. 2021, 22, 5830. [Google Scholar] [CrossRef] [PubMed]
- Wang, S.; Meckling, K.A.; Marcone, M.F.; Kakuda, Y.; Tsao, R. Synergistic, Additive, and Antagonistic Effects of Food Mixtures on Total Antioxidant Capacities. J. Agric. Food Chem. 2011, 59, 960–968. [Google Scholar] [CrossRef] [PubMed]
- Yang, T.W.; Wang, C.C.; Sung, W.W.; Ting, W.C.; Lin, C.C.; Tsai, M.C. The Effect of Coffee/Caffeine on Postoperative Ileus Following Elective Colorectal Surgery: A Meta-Analysis of Randomized Controlled Trials. Int. J. Color. Dis. 2022, 37, 623–630. [Google Scholar] [CrossRef]
- Nasi, M.; De Gaetano, A.; Carnevale, G.; Bertoni, L.; Selleri, V.; Zanini, G.; Pisciotta, A.; Caramaschi, S.; Reggiani Bonetti, L.; Farinetti, A.; et al. Effects of Energy Drink Acute Assumption in Gastrointestinal Tract of Rats. Nutrients 2022, 14, 1928. [Google Scholar] [CrossRef]
- Gallego-Barceló, P.; Bagues, A.; Benítez-Álvarez, D.; López-Tofiño, Y.; Gálvez-Robleño, C.; López-Gómez, L.; Dolores, M.; Castillo, D.; Abalo, R. Evaluation of the Effects of Instant Cascara Beverage on the Brain-Gut Axis of Healthy Male and Female Rats. Nutrients 2023, 16, 65. [Google Scholar] [CrossRef]
- Del Castillo, M.D.; Ibáñez, E.; Amigo-Benavent, M.; Herrero, M.; Plaza, M.; Ullate, M. Application of Products of Coffee Silverskin in Anti-Ageing Cosmetics and Functional Food. WO2013004873A1, 10 January 2013. [Google Scholar]
- Iriondo-Dehond, A.; Iriondo-Dehond, M.; Del Castillo, M.D. Applications of Compounds from Coffee Processing By-Products. Biomolecules 2020, 10, 1219. [Google Scholar] [CrossRef]
- López-Gómez, L.; López-Tofiño, Y.; Abalo, R. Dependency on Sex and Stimulus Quality of Nociceptive Behavior in a Conscious Visceral Pain Rat Model. Neurosci. Lett. 2021, 746, 135667. [Google Scholar] [CrossRef]
- Cora, M.C.; Kooistra, L.; Travlos, G. Vaginal Cytology of the Laboratory Rat and Mouse:Review and Criteria for the Staging of the Estrous Cycle Using Stained Vaginal Smears. Toxicol. Pathol. 2015, 43, 776–793. [Google Scholar] [CrossRef]
- Ranganathan, P.; Jayakumar, C.; Manicassamy, S.; Ramesh, G. CXCR2 Knockout Mice Are Protected against DSS-Colitis-Induced Acute Kidney Injury and Inflammation. Am. J. Physiol. Ren. Physiol. 2013, 305, 1422–1427. [Google Scholar] [CrossRef]
- Uranga, J.A.; García-Martínez, J.M.; García-Jiménez, C.; Vera, G.; Martín-Fontelles, M.I.; Abalo, R. Alterations in the Small Intestinal Wall and Motor Function after Repeated Cisplatin in Rat. Neurogastroenterol. Motil. 2017, 29, e13047. [Google Scholar] [CrossRef]
- López-Tofiño, Y.; Barragán del Caz, L.F.; Benítez-Álvarez, D.; Molero-Mateo, P.; Nurgali, K.; Vera, G.; Bagües, A.; Abalo, R. Contractility of Isolated Colonic Smooth Muscle Strips from Rats Treated with Cancer Chemotherapy: Differential Effects of Cisplatin and Vincristine. Front. Neurosci. 2023, 17, 1304609. [Google Scholar] [CrossRef] [PubMed]
- Straface, M.; Makwana, R.; Palmer, A.; Rombolà, L.; Aleong, J.C.; Morrone, L.A.; Sanger, G.J. Inhibition of Neuromuscular Contractions of Human and Rat Colon by Bergamot Essential Oil and Linalool: Evidence to Support a Therapeutic Action. Nutrients 2020, 12, 1381. [Google Scholar] [CrossRef] [PubMed]
- Vera, G.; Castillo, M.; Cabezos, P.A.; Chiarlone, A.; Martín, M.I.; Gori, A.; Pasquinelli, G.; Barbara, G.; Stanghellini, V.; Corinaldesi, R.; et al. Enteric Neuropathy Evoked by Repeated Cisplatin in the Rat. Neurogastroenterol. Motil. 2011, 23, 370-e163. [Google Scholar] [CrossRef] [PubMed]
- Abalo, R.; Rivera, A.J.; Vera, G.; Martín, M.I. Ileal Myenteric Plexus in Aged Guinea-Pigs: Loss of Structure and Calretinin-Immunoreactive Neurones. Neurogastroenterol. Motil. 2005, 17, 123–132. [Google Scholar] [CrossRef]
- Chen, Y.; Kim, M.; Paye, S.; Benayoun, B.A. Sex as a Biological Variable in Nutrition Research: From Human Studies to Animal Models. Annu. Rev. Nutr. 2022, 42, 227–250. [Google Scholar] [CrossRef] [PubMed]
- Thein, W.; Po, W.W.; Kim, D.M.; Sohn, U.D. The Altered Signaling on EFS-Induced Colon Contractility in Diabetic Rats. Biomol. Ther. 2020, 28, 328. [Google Scholar] [CrossRef] [PubMed]
- Sánchez, M.; Suárez, L.; Andrés, M.T.; Flórez, B.H.; Bordallo, J.; Riestra, S.; Cantabrana, B. Modulatory Effect of Intestinal Polyamines and Trace Amines on the Spontaneous Phasic Contractions of the Isolated Ileum and Colon Rings of Mice. Food Nutr. Res. 2017, 61, 1321948. [Google Scholar] [CrossRef] [PubMed]
- Ma, Y.; Qiao, X.; Falone, A.E.; Reslan, O.M.; Sheppard, S.J.; Khalil, R.A. Gender-Specific Reduction in Contraction Is Associated with Increased Estrogen Receptor Expression in Single Vascular Smooth Muscle Cells of Female Rat. Cell. Physiol. Biochem. 2010, 26, 457–470. [Google Scholar] [CrossRef] [PubMed]
- Al-Shboul, O.A.; Al-Dwairi, A.N.; Alqudah, M.A.; Mustafa, A.G. Gender Differences in the Regulation of MLC20 Phosphorylation and Smooth Muscle Contraction in Rat Stomach. Biomed. Rep. 2018, 8, 283. [Google Scholar] [CrossRef] [PubMed]
- Tokutomi, Y.; Tokutomi, N.; Nishi, K. The Properties of Ryanodine-Sensitive Ca2+ Release in Mouse Gastric Smooth Muscle Cells. Br. J. Pharmacol. 2001, 133, 125. [Google Scholar] [CrossRef] [PubMed]
- Tazzeo, T.; Bates, G.; Roman, H.N.; Lauzon, A.M.; Khasnis, M.D.; Eto, M.; Janssen, L.J. Caffeine Relaxes Smooth Muscle through Actin Depolymerization. Am. J. Physiol. Lung Cell Mol. Physiol. 2012, 303, L334. [Google Scholar] [CrossRef] [PubMed]
- Lee, H.T.; Hennig, G.W.; Fleming, N.W.; Keef, K.D.; Spencer, N.J.; Ward, S.M.; Sanders, K.M.; Smith, T.K. The Mechanism and Spread of Pacemaker Activity through Myenteric Interstitial Cells of Cajal in Human Small Intestine. Gastroenterology 2007, 132, 1852–1865. [Google Scholar] [CrossRef]
- Ratz, P.H.; Berg, K.M.; Urban, N.H.; Miner, A.S. Regulation of Smooth Muscle Calcium Sensitivity: KCl as a Calcium-Sensitizing Stimulus. Am. J. Physiol. Cell Physiol. 2005, 288, C769–C783. [Google Scholar] [CrossRef]
- Montgomery, L.E.A.; Tansey, E.A.; Johnson, C.D.; Roe, S.M.; Quinn, J.G. Autonomic Modification of Intestinal Smooth Muscle Contractility. Adv. Physiol. Educ. 2016, 40, 104–109. [Google Scholar] [CrossRef]
- Zhou, H.; Kong, D.H.; Pan, Q.W.; Wang, H.H. Sources of Calcium in Agonist-Induced Contraction of Rat Distal Colon Smooth Muscle in Vitro. World J. Gastroenterol. 2008, 14, 1077–1083. [Google Scholar] [CrossRef] [PubMed]
- Broad, J.; Kung, V.W.S.; Palmer, A.; Elahi, S.; Karami, A.; Darreh-Shori, T.; Ahmed, S.; Thaha, M.A.; Carroll, R.; Chin-Aleong, J.; et al. Changes in Neuromuscular Structure and Functions of Human Colon during Ageing Are Region-Dependent. Gut 2019, 68, 1210–1223. [Google Scholar] [CrossRef] [PubMed]
- Hegde, S.; Shi, D.W.; Johnson, J.C.; Geesala, R.; Zhang, K.; Lin, Y.M.; Shi, X.Z. Mechanistic Study of Coffee Effects on Gut Microbiota and Motility in Rats. Nutrients 2022, 14, 4877. [Google Scholar] [CrossRef] [PubMed]
- Aulí, M.; Martínez, E.; Gallego, D.; Opazo, A.; Espín, F.; Martí-Gallostra, M.; Jiménez, M.; Clavé, P. Effects of Excitatory and Inhibitory Neurotransmission on Motor Patterns of Human Sigmoid Colon in Vitro. Br. J. Pharmacol. 2008, 155, 1043. [Google Scholar] [CrossRef] [PubMed]
- Sanger, G.J.; Broad, J.; Kung, V.; Knowles, C.H. Translational Neuropharmacology: The Use of Human Isolated Gastrointestinal Tissues. Br. J. Pharmacol. 2013, 168, 28–43. [Google Scholar] [CrossRef] [PubMed]
- Yamato, S.; Kurematsu, A.; Amano, T.; Ariga, H.; Ando, T.; Komaki, G.; Wada, K. Urocortin 1: A Putative Excitatory Neurotransmitter in the Enteric Nervous System. Neurogastroenterol. Motil. 2020, 32, e13842. [Google Scholar] [CrossRef] [PubMed]
- Hinds, N.M.; Ullrich, K.; Smid, S.D. Cannabinoid 1 (CB1) Receptors Coupled to Cholinergic Motorneurones Inhibit Neurogenic Circular Muscle Contractility in the Human Colon. Br. J. Pharmacol. 2006, 148, 191–199. [Google Scholar] [CrossRef] [PubMed]
- Broad, J.; Mukherjee, S.; Samadi, M.; Martin, J.E.; Dukes, G.E.; Sanger, G.J. Regional- and Agonist-Dependent Facilitation of Human Neurogastrointestinal Functions by Motilin Receptor Agonists. Br. J. Pharmacol. 2012, 167, 763–774. [Google Scholar] [CrossRef] [PubMed]
- Li, C.; Micci, M.A.; Murthy, K.S.; Pasricha, P.J. Substance P Is Essential for Maintaining Gut Muscle Contractility: A Novel Role for Coneurotransmission Revealed by Botulinum Toxin. Am. J. Physiol. Gastrointest. Liver Physiol. 2014, 306, G839. [Google Scholar] [CrossRef]
- Drimousis, S.; Markus, I.; Murphy, T.v.; Shevy Perera, D.; Phan-Thien, K.C.; Zhang, L.; Liu, L. Gender-Related Differences of Tachykinin NK 2 Receptor Expression and Activity in Human Colonic Smooth Muscle. J. Pharmacol. Exp. Ther. 2020, 375, 28–39. [Google Scholar] [CrossRef]
- Di Natale, M.R.; Hunne, B.; Liew, J.J.M.; Fothergill, L.J.; Stebbing, M.J.; Furness, J.B. Morphologies, Dimensions and Targets of Gastric Nitric Oxide Synthase Neurons. Cell Tissue Res. 2022, 388, 19. [Google Scholar] [CrossRef]
- Idrizaj, E.; Traini, C.; Vannucchi, M.G.; Baccari, M.C. Nitric Oxide: From Gastric Motility to Gastric Dysmotility. Int. J. Mol. Sci. 2021, 22, 9990. [Google Scholar] [CrossRef]
- Li, Q.; Zhao, B.; Li, W.; He, Y.; Tang, X.; Zhang, T.; Zhong, Z.; Pan, Q.; Zhang, Y. Effects of repeated drug administration on behaviors in normal mice and fluoxetine efficacy in chronic unpredictable mild stress mice. Biochem. Biophys. Res. Commun. 2022, 615, 36–42. [Google Scholar] [CrossRef]
- Hoggatt, A.F.; Hoggatt, J.; Honerlaw, M.; Pelus, L.M. A Spoonful of Sugar Helps the Medicine Go Down: A Novel Technique to Improve Oral Gavage in Mice. J. Am. Assoc. Lab. Anim. Sci. 2010, 49, 329–334. [Google Scholar]
- McDonnell-Dowling, K.; Kleefeld, S.; Kelly, J.P. Consequences of Oral Gavage during Gestation and Lactation on Rat Dams and the Neurodevelopment and Behavior of Their Offspring. J. Am. Assoc. Lab. Anim. Sci. 2017, 56, 79. [Google Scholar]
- Gil, M.C.; Aguirre, J.A.; Lemoine, A.P.; Segura, E.T.; Barontini, M.; Armando, I. Influence of Age on Stress Responses to Metabolic Cage Housing in Rats. Cell. Mol. Neurobiol. 1999, 19, 625–633. [Google Scholar] [CrossRef]
- Labanski, A.; Langhorst, J.; Engler, H.; Elsenbruch, S. Stress and the Brain-Gut Axis in Functional and Chronic-Inflammatory Gastrointestinal Diseases: A Transdisciplinary Challenge. Psychoneuroendocrinology 2020, 111, 104501. [Google Scholar] [CrossRef]
- Bagues, A.; Lopez-Tofiño, Y.; Galvez-Robleño, C.; Abalo, R. Effects of Two Different Acute and Subchronic Stressors on Gastrointestinal Transit in the Rat: A Radiographic Analysis. Neurogastroenterol. Motil. 2021, 33, e14232. [Google Scholar] [CrossRef]
- Gálvez-Robleño, C.; López-Tofiño, Y.; López-Gómez, L.; Bagüés, A.; Soto-Montenegro, M.L.; Abalo, R. Radiographic Assessment of the Impact of Sex and the Circadian Rhythm-Dependent Behaviour on Gastrointestinal Transit in the Rat. Lab. Anim. 2023, 57, 270–282. [Google Scholar] [CrossRef]
- Chey, W.D.; Hashash, J.G.; Manning, L.; Chang, L. AGA Clinical Practice Update on the Role of Diet in Irritable Bowel Syndrome: Expert Review. Gastroenterology 2022, 162, 1737–1745.e5. [Google Scholar] [CrossRef]
Components | Instant Cascara Beverage (10 mg/mL) |
---|---|
Carbohydrates (mg/mL) | 4.748 |
Total fiber (mg/mL) | 1.832 |
Lipids (mg/mL) | 0.058 |
Protein (mg/mL) | 0.625 |
Magnesium (mg/mL) | 2.084 × 10−3 |
Sodium (mg/mL) | 26.658 × 10−3 |
Potassium (mg/mL) | 228.4 × 10−3 |
Calcium (mg/mL) | 5.478 × 10−3 |
Caffeine (mg/mL) | 0.139 |
Phenolic content (mg eq. Cga/mL) | 0.89 |
Chlorogenic acids (mg/mL) | 1.07–1.26 |
Melanoidins (mg/mL) | 1.5 |
Males—Control | Males—IC | Females—Control | Females—IC | ||||||
---|---|---|---|---|---|---|---|---|---|
Initial (g) | Final (g) | Initial (g) | Final (g) | Initial (g) | Final (g) | Initial (g) | Final (g) | ||
LM | PA | 0.40 ± 0.07 | 0.35 ± 0.06 | 0.38 ± 0.06 | 0.31 ± 0.05 | 0.41 ± 0.05 | 0.33 ± 0.05 | 0.44 ± 0.05 | 0.36 ± 0.05 |
TA | 0.24 ± 0.04 | 0.21 ± 0.05 | 0.27 ± 0.05 | 0.18 ± 0.03 | 0.18 ± 0.03 | 0.19 ± 0.05 | 0.22 ± 0.03 | 0.19 ± 0.03 | |
CM | PA | 1.73 ± 0.21 | 1.53 ± 0.19 | 1.47 ± 0.18 | 1.48 ± 0.18 | 0.96 ± 0.13 ## | 0.96 ± 0.10 # | 1.14 ± 0.09 | 1.31 ± 0.08 |
TA | 0.75 ± 0.11 | 0.70 ± 0.12 | 0.80 ± 0.11 | 0.63 ± 0.11 | 0.55 ± 0.09 # | 0.37 ± 0.05 | 0.74 ± 0.09 | 0.54 ± 0.06 |
Antibody | Parameter (Unit) | Males—Control | Males—IC | Females—Control | Females—IC |
---|---|---|---|---|---|
HuC/D | Ganglionic area (%) | 10.90 ± 1.21 | 10.59 ± 1.28 | 10.95 ± 0.59 | 11.05 ± 1.28 |
Ganglionic size (μm2) | 25,716 ± 2690 | 22,345 ± 2750 | 20,332 ± 3277 | 25,126 ± 3199 | |
Neurons/ganglion | 54.06 ± 4.933 | 47 ± 4.77 | 41.91 ± 9.12 | 49.77 ± 6.431 | |
Total neuronal density (neurons/mm2) | 231.40 ±19.46 | 212 ± 17.64 | 217.40 ± 23.49 | 219.90 ± 28.79 | |
Packing density (intraganglionic neurons/ganglionic area) | 2156 ± 226.1 | 2147 ± 134.7 | 2000 ± 143.1 | 1976 ± 45.15 | |
Extraganglionic neuronal density (extraganglionic neurons/serosal area) | 3.75 ± 0.78 | 4.28 ± 0.89 | 4.81 ± 0.35 | 3.03 ± 1.026 | |
nNOS | Inhibitory neurons (%) | 18.67 ± 2.24 | 22.13 ± 2.36 | 18.93 ± 2.02 | 22.25 ± 4.02 |
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content. |
© 2024 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Gallego-Barceló, P.; Benítez-Álvarez, D.; Bagues, A.; Silván-Ros, B.; Montalbán-Rodríguez, A.; López-Gómez, L.; Vera, G.; del Castillo, M.D.; Uranga, J.A.; Abalo, R. Ex Vivo Study of Colon Health, Contractility and Innervation in Male and Female Rats after Regular Exposure to Instant Cascara Beverage. Foods 2024, 13, 2474. https://doi.org/10.3390/foods13162474
Gallego-Barceló P, Benítez-Álvarez D, Bagues A, Silván-Ros B, Montalbán-Rodríguez A, López-Gómez L, Vera G, del Castillo MD, Uranga JA, Abalo R. Ex Vivo Study of Colon Health, Contractility and Innervation in Male and Female Rats after Regular Exposure to Instant Cascara Beverage. Foods. 2024; 13(16):2474. https://doi.org/10.3390/foods13162474
Chicago/Turabian StyleGallego-Barceló, Paula, David Benítez-Álvarez, Ana Bagues, Blanca Silván-Ros, Alba Montalbán-Rodríguez, Laura López-Gómez, Gema Vera, María Dolores del Castillo, José A. Uranga, and Raquel Abalo. 2024. "Ex Vivo Study of Colon Health, Contractility and Innervation in Male and Female Rats after Regular Exposure to Instant Cascara Beverage" Foods 13, no. 16: 2474. https://doi.org/10.3390/foods13162474